CA2122592C - System for delivering an active substance for sustained release - Google Patents

Abstract

A system for delivering an active substance has sustained release of the active substance in the intestinal tract. This delivery system is especially useful for an active substance such as a .beta.-lactam antibiotic which preferably has minimal exposure to the acidic environment of the stomach. Particles comprise an active ingredient disposed in a core which has at least one coating of a prolamine and one coating of an enteric compound thereon. The particles may be very small and suspended in a liquid medium.

Description

2 PCI`/US92/11160 `~ ~122~92 SYSTEM FOR DELIVERING AN ACTIVE SUBSTANCE FOR SUSTAINED RELEASE
The present invention relates generally to systems for sustained release of active substances in the digestive tract and more specifically to microcapsules in suspensions as a system for delivering an active substance for sustained release in the intestinal tract.

Some medical conditions are best treated by administration of a pharmaceutical or other active substance which is formulated to allow the active substance or ingredient to act as quickly as possible. Such a formulation may comprise an injectable solution, suspension, or a readily dissolvable tablet or capsule. This type of formulation is useful, for instance, for treating acute pain, such as headaches, or pain associated with sudden trauma, such as an accident.
Other medical conditions are best treated by administration of a pharmaceutical or other active substance in such a way as to sustain its action over an extended period of time. This type of administration is useful, for example, for treating chronic pain, such as that associated with rheumatic or arthritic conditions, for the treatment of a chronic cardiovascular condition, or for administering an antibiotic in a course of treatment covering several days. Sustained action can be achieved by - repeated administration of an immediate-release tablet or capsule at frequent intervals, for instance every four hours. However, this is general1y inconvenient, especially during the night, when it is often necessary to awaken a patient to administer the suspension, tablet or capsule. In addition, such multiple dosing may lead to undesirable fluctuations in the plasma concentration of the active substance.
It has previously been proposed to produce a formulation which will release the active substance therein at a controlled rate such that the amount available in the body to treat the condition is maintained at a therapeutic level over an extended period of time. Particularly suitable periods are twelve hours and twenty-four hours, since such formulations need only be taken once or twice a day to maintain an effective treatment of the condition. Such formulations are generally known as "sustained-release formulations."
Many sustained-release formulations are already known, but there is W O 93/12772 2122~92 P~r/u592/lll6o no generally applicable method by which such formulations can be designed.
Generally speaking, each sustained-released formulation is dependent on the particular active substance incorporated therein. In designing a formulation, it is generally necessary to take into account many factors, including the rates of absorption and clearance of the active substance, the interaction of the active substance with the excipients and/or coating to be used in the formulation, the solubility of the active substance and of the excipients and/or coatings, and the effects on the bioavailability of the active substance which may be caused by the excipients and/or coatings.
It is, however, not possible to readily predict whether any particular formulation will provide the desired sustained-release, and it is generally found necessary to carry out considerable experimentation to produce a sustained-release formulation having the desired properties.
The challenge of providing a sustained release delivery system is greatly increased when the patient is an infant, young child, or a more mature person who is unable to easily ingest large tablets or capsules.
Such persons are more amenable to the ingestion of pharmaceutical or other active substances via liquid suspensions. The challenge is increased exponentially when for such persons the active compound, which is to be delivered, such as a ~-lactim antibiotic, is most effective when protected from acidic gastric juices and is desired to be gradually released in the intestine. These challenges are met by the system for delivery of an active substance which is disclosed herein.

DESCRIPTION OF THE PRIOR ART
The encapsulation of active substance is well known for a variety of purposes, among them to protect the active substance from degradation when in contact with other agents in a given product or composition; to modulate the release of the active substance, and to render the active substance capable of withstanding rigorous processing conditions during formulation into products.
Numerous prior art patents and publications address the objective of providing a sustained release formulation of an active substance in the gastrointestinal tract.
U.S. Patent 4,525,339 teaches the desirability of protecting a ~-lactim antibiotic from exposure to gastric juices, this being achieved by W O 93/12772 pc~r/uss2/l1l6o 2122~92 an enteric coating. One of the possible enteric coating materials suggested in this patent is zein. However; there is no suggestion of using zein in combination with another enteric coating. Furthermore, the data set forth below in the present document, (Figure 1), indicates that zein is not a suitable material for use as an enteric coating unless an undesirably thick coating layer is used.
U.S. Patent 4,079,131 teaches a liquid suspension for providing amoxicillin to a patient, but it does not address the issue of sustained release. The amoxicillin is suspended in an anhydrous vegetable oil vehicle containing a saccharide. However; an oil based suspension has taste characteristics which may severely impair the ease of administering amoxicillin to a young child. Additionally this patent does not address sustained release through use of a water based suspension.
Japanese Kokoku Sho 45-12759 teaches that a mixture of 80-99% zein and 1-20% HPMC (3-15% hydroxypropoxy group and 19-32% methoxy group) may be used to coat tablets for taste masking, but the resulting tablets are readily dissolved in gastric juice. This patent reference teaches that hydroxy propyl methyl cellulose must be mixed with zein to obtain taste mask;ng properties.
~ Pharmacokinetics and bioavailability of a controlled release amoxicillin formulation", Arancibia et al, INTERNATIONAL JOURNAL OF CLINICAL
PHARMACOLOGY THERAPY AND TOXICOLOGY, Volume 25 No. 2, 1987, pages 97-100, reports the evaluation of an unsatisfactory sustained release system (which is not described) in an in vivo experiment. This article indicates that the sustained release system did, however, perform satisfactorily in-vitro dissolution studies. "Biopharmaceutical Evaluation of Sustained-Release Ethylcellulose Microcapsules Containing Amoxicillin Using Beagle Dogs", Uchida et al., CHEMICAL PHARMACEUTICAL BULLETIN, Volume 37 No. 12, (1989), pages 3416-3419 shows that the desirability of a sustained-release amoxicillin has been recognized for some time, but this article does not address the desirability of delaying release of the amoxicillin until the drug is in the intestine.
U.S. Patents 4,876,097 and 4,983,403 teach that zein may be used in a coating material that will release the core substance at pH's of < 3.5, but will be stable in a pH of > 5. These patents teach the use of such a delivery system for delivering an active substance to the second stomach of WO 93/12772 2 2 5 ~ 2 PCI`/US92/11160 a ruminant, but do not address the issue of sustained release. However, the data presented in the present patent application does not support these pH
ranges for the zein used in practicing the present invention.
Japanese Koka Sho 61-141862 teaches coating vitamin C cores with zein or shellac to obtain time release, but concludes that shellac is superior to zein for this purpose. The exact type of zein used in the experiments is not disclosed.
Japanese Kokoku Hei 3-988 teaches that a health food which is either inside of a gelatin capsule or coated with gelatin may advantageously be overcoated with a "high quality zein" to prevent the health food from being released until reaching the pylorus of the stomach. However, there is no teaching or indication that sustained release of the health food in the intestine is achievable with this method, or even is desirable. The main objective is to prevent the health food from being exposed in the mouth.
British Patent 935,672 (published September 4, 1963) teaches a "sustained release tablet" having an active substance dispersed in a matrix which contains zein. However, the structure disclosed in this patent gives a big initial release in gastric fluid, as opposed to the present invention, followed by a much slower release rate in intestinal fluid.
U.S. Patent 3,$58,768 teaches sustained release pharmaceutical compositions with a core of an active compound dispersed in a matrix which contains zein, with the core being coated with a layer of the active substance dispersed in a matrix containing a hydrophilic gum. The test data presented in this patent shows release of the active substance in gastric fluid to be substantially the same as in intestinal fluid.
U.S. Patent 2,895,880 teaches that an active compound may be dispersed in a matrix containing zein in order to achieve time release of the active compound when orally ingested. However, there is no teaching or suggestion in this patent that release of the active compound in the stomach should be minimized. However, the data presented in the present application shows that a minimal release of an active substance in gastric juices is not achieved by such a structure.
European Patent Application 0130387 (published March 21, 1984) teaches the use of a "lower level" of zein in the matrix of a tablet, and the zein is not used as a coating material. This tablet releases a large burst dosage in acid, followed by slower sustained release in a base. However, W O 93~12772 pc~r/us92/ll16o this is not the release pattern desired to be achieved in the present invention.
U.S. Patent 4,892,742 teaches an active substance in a matrix which contains zein, and the core is coated with a "rate controlling polymer".
U.S. Patent 3,802,896 teaches a coating solution which contains zein and has utility for taste masking, and possible sustained release, of an active substance. However, the use of only a single layer of this coating solution, not in combination with a layer of any other material, is disclosed in this patent.
U.S. Patent 3,939,259 teaches a sustained release system of gelatin capsules containing: (a) uncoated particles of an active substance to be released in the stomach during the first hour after ingestion; (b) particles of the active substance coated with a mixture of zein and shellac to be released two hours after ingestion; and particles of the active substance with a thicker coating of the zein and shellac mixture which is to be released four hours after ingestion. This sustained release system is not designed to minimize release of the active ingredient in the stomach.
Japanese Kokai Sho 62-201823 teaches that beneficial bifido bacteria and lactic acid bacteria may be delivered to the intestine, and protected from exposure to gastric juices, by overcoating the capsule, or microcapsule, which contains the bacteria with zein. However, this structure readily underwent disintegration in intestinal juices within 3-12 minutes, which does not solve the problem of providing sustained release in the intestine.
U.S. Patent 4,308,251 teaches a twice-a-day sustained release aspirin tablet having both a time release controlling agent, (preferably cellulose acetate phthalate, but could be zein), and an erosion promoting agent such as corn starch. However, there is no teaching or suggestion that an enteric coating should be used to minimize release of the aspirin in the stomach.
U.S. Patent 4,137,300 teaches a sustained release dosage system having: (a) a core comprising an active substance and at least two members ~ selected from the group consisting of a higher alkanol and alkanoic acid;
and (b) a coating of a prolamine overlying the core. However; no time-release data is presented in this patent, and there is no teaching orsuggestion to use an enteric coating in combination with the prolamine coating.

WO 93/12772 PCI`/US92/11160 U.S. Patent 4,876,094 teaches a controlled release liquid dosage formulation wherein a core of an active substance has a first coating of a fat which melts at less than 38.3OC, and is overcoated with cellulose acetate or zein. The carrier liquid has a pH of less than 5 and comprises a viscous solution of a sugar, preferably being a high fructose corn syrup.
The above noted and other known techniques have heretofore provided no solution for the development of a delivery system for active substances which remedies the aforenoted problems by permitting the sustained release of an active substance in the intestine with little or no release in the stomach.

In the accompanying drawings:
Figure 1 is a graph showing the dissolution of acetaminophen from the cores of particles having a single zein coating thereon at a variety of weight percentages of the core, when the particles were placed in simulated gastric fluid at pH 1.2;

Figure 2 is a graph showing the dissolution of acetaminophen from the cores of particles having a single zein coating thereon at various weight percentages of the core, when the particles were placed in simulated intestinal fluid at pH 6.8;

Figure 3 is a graph showing the dissolution of acetaminophen from the cores of particles having a zein coating over the core, and an enteric coating over the zein coating, when the particles were placed in simulated gastric fluid at pH 1.2;

Figure 4 is a graph showing the dissolution of acetaminophen from the cores of particles having a zein coating over the core and an enteric coating over the zein coating, when the particles were placed in simulated intestinal fluid at pH 6.8;

Figure 5 is a graph showing the dissolution of acetaminophen from the cores of particles haying a first zein coating over the core, with an enteric coat;ng over the first zein coating, followed by a second coating WO 93~12772 PCI`/US92/11160 2l22~92 of zein over the enteric coating, the particles were placed in simulated gastric fluid at pH 1.2;

Figure 6 is a graph showing the dissolution of acetaminophen from the cores of particles having a first zein coating over the core, with an enteric coating over the first zein coating, followed by a second coating of zein over the enteric coating, when the particles were placed in simulated intestinal fluid at pH 6.8;

Figure 7 is a graph showing the dissolution of acetaminophen from the cores of particles having a first zein coating over the core, with an enteric coating over the first zein coating, followed by a second coating of zein over the enteric coating, when the particles were sequentially placed in simulated gastric fluid at pH 1.2 for one hour, followed by placement in simulated intestinal fluid at pH 6.8 for six hours;

Figure 8 is a graph showing the dissolution of amoxicillin from the cores of particles having a zein coating over the core and an enteric coating over the zein coating, when the particles were placed in simulated gastric fluid at pH 1.2;

Figure 9 is a graph showing the dissolution of amoxicillin from the cores of particles having a zein coating over the core and a second enteric coating over the zein coating, when the particles were placed in simulated intestinal fluid at pH 6.8; and Figure 10 is a graph showing the dissolution of amoxicillin from the cores of particles having a first zein coating over the core, with a second enteric coating over the first zein coating, followed by a second coating of zein over the enteric coating, when the particles were sequentially placed in simulated gastric fluid at pH 1.2 for one hour, followed by ~ placement in simulated intestinal fluid at pH 6.8 for six hours.

- DETAILED DESCRIPTION OF THE INVENTION
A delivery system for active substances is produced by a process which comprlses formlng the active compound and optionally one or more excipients WO 93/12772 PCI`/US92/11160 and binders into a core, preferably using a rotor insert with fluid bed coating; applying a first coating to this core, preferably by fluid bed coating and applying a second coating over the particles thus formed, preferably by fluid bed coating. In some embodiments a third coating is then applied over the particles thus formed, preferably using fluid bed coating. It is preferable to screen the core particles initially formed before applying the first coating to enhance uniformity in the final product.
The active substance delivery system may be incorporated into a variety of pharmaceutical and nutritional products including pharmaceutical suspensions, pediatric infant nutritional formulas, and nutritional preparations. The present invention therefore encompasses pharmaceutical suspensions, pediatric infant formulas, and nutritional preparations, such as medical nutritionals for general health, as well as disease specific medical nutritionals, all incorporating the present active compound delivery system.
In accordance with the present invention, a delivery system for active substances is disclosed which comprises a composite particle structure having minimal dissolution in a suspension, minimal release in mouth and stomach of an active substance, and sustained release of the active substance in the intestinal tract. The present invention is a two or three coating layer encapsulation system for delivering an active substance to the intestine where the active substance will be slowly released.
A delivery system for an active substance in accordance with a first double coated embodiment of the invention generally comprises:
(a) a core comprising an active substance, or an active substance in a matrix with excipients;
(b) a first coating on the core comprising a prolamine in an amount from about 10% to about 70%, preferably about 20% to 50%, by weight of the total weight of the core; and (c) a second coating overlying the first coat and comprising at least one enteric compound in an amount from about lOYo to about 707O~
preferably about 20% to 60%, by weight of the sum of the core material and the first coating.

WO 93/12772 PCI`/US92/11160 2122~92 g A delivery system for an active substance in accordance with a second double coated embodiment of the invention generally comprises:
(a) a core comprising an active substance, or an active substance in a matrix with excipients;
(b) a first coating on the core comprising at least one enteric compound in an amount from about 10% to about 70%, preferably about 20% to 40%~ by weight of the total weight of the core; and (c) a second coating overlying the first coat and comprising a prolamine in an amount of from about 20% to 100%, preferably about 4Wo to 60%, by weight of the sum of the weights of the core and the first coating.

A delivery system for an active substance in accordance with a triple coated embodiment of with the present invention generally comprises:
(a) a core comprising an active substance, or an active substance in a matrix with excipients;
(b) a first coating comprising a prolamine in an amount from about 10%
to about 70%, preferably about 10% to 30%, by weight of the total weight of the core;
(c) a second coating overlying the first coat and comprising at least one enteric compound in an amount from about 5% to about 70YO~
preferably about 20% to 40Z~ by weight of the sum of the total weights of the core and the first coating; and (d) a third coating comprising a prolamine in an amount of from about 20% to 100%, preferably about 40% to 70%, by weight of the sum of the weights of the core and the first two coatings.

Prolamines form the main protein components of cereal grains and flour. Unlike all other proteins, they can be extracted from flour with 80 percent alcohol, but they are insoluble in absolute alcohol and water. The most important prolamines are zein, gliadin, and hordein. Zein is preferred in the present invention.
As used herein and in the claims an "enteric compound" is understood - to mean a composition of matter which is generally resistant to disintegration in human gastric fluids, but will disintegrate in human intestinal fluids, as well as compositions of matter which disintegrate very WO 93~12772 212 2 5 9 2 PCr/US92/1 1160 slowly in human gastric fluids, but more rapidly in human intestinal fluids.
The active substances which are believed to be suitable for incorporation in the core of an encapsulated particle in accordance with the present invention are bioactive substances including for example, analgesics, antibiotics, oncolytics, immunogens, antidepressants and other psychotherapy drugs, antivirals, drugs to treat HIV compromised individuals and individuals with AIDS, immuno-modulators, vitamins, dietary fiber and other nutrients, enzymes, hormones, vaccines, antibodies and other pharmaceuticals. The foregoing list is not intended to be inclusive but merely representative of various active compounds both simple and complex that are contemplated in accordance with the present invention.
A core containing an active substance(s) is prepared by standard processes such as spray drying, fluid bed coating, fluid bed coating with a rotor insert, and may optionally be prepared with additives such as excipients, including bulking agents, fillers, and binders. The excipients are generally present in amounts of up to 80% by weight of the total core material and can be mixed in combination with each other or used individually. Suitable excipients include, but are not limited to, carbohydrate materials, polyhydric alcohols, binders that are soluble at a pH greater than about 5.5, and mixtures thereof. Carbohydrates useful as excipients include traditional water-soluble sweetening agents such as monosaccharides, disaccharides and polysaccharides such as xylose, ribose, glucose, lactose, mannose, galactose, fructose, dextrose, sucrose, sugar, maltose, partially hydrolyzed starch, or corn syrup solids and sugar alcohols such as sorbitol, xylitol, mannitol and the like, and mixtures thereof. Suitable binders include for example zein, polyvinylpyrrolidone, also known as PVP, methacrylic acid copolymer USP/NF Type A, methacrylic acid copolymer USP/NF Type B, methacrylic acid copolymer USP/NF Type C, blends of methacrylic acid copolymer USP/NF Type A and methacrylic acid copolymer USP/NF Type B, hydroxy propyl methyl cellulose phthalate, also known as HPMCP, HP50 or HP55, cellulose acetate phthalate, also known as C-A-P, cellulose acetate trimellitate, also known as C-A-T, blends of C-A-T
and C-A-P, ethyl cellulose, and polyvinyl acetate phthalate, also known as PVAP. Gums, pectins, aliginates, mucilages, and mixtures thereof may also serve as suitable binders. Suitable binders in this group include gum arabic, tragacanth, karaya, ghattiagar, aliginates, carrageenans, .

fuercellaran, psyllium, and mixtures thereof.
Coating layers of the present invention which are not intended to have enteric characteristics comprise zein or other prolamines and a plasticizer or hydrophobic substance, or optionally, both a plasticizer and hydrophobic substance. The zein component for a non-enteric first coating layer preferably comprises zein with an ash content of 2% or less by weight. The method used to determine ash is in the USP XXII, "Residue on Ignition", sulfated. The zein used in many of the examples set forth herein was F
4000, manufactured by Freeman Industries, Tuckahoe, New York, U.S.A., with an ash content of about 1.1% by weight and zein F-400-LE from Freeman Industries with an ash content of about 0.07% by weight. The plasticizer may be generally selected from the group consisting of food grade glycols including triethylene glycol and propylene glycol, acetylated glycerides, oleic acid, lactic acid acetamide, ethylene glycol monooleate, glycerin, glycerol monostearate, dibutyl tartrate, and tricresol phosphate. A
suitable hydrophobic substance used for the zein coating material comprises vegetable and animal fats, either unhydrogenated, hydrogenated, or partially hydrogenated, fatty acids, and glycerine esters of fatty acids, with representative materials comprising palm oil, palm kernel oil, soybean oil, rapeseed oil, rice bran oil, sunflower oil, safflower oil, coconut oil, castor oil, MCT oil, also known as glycerine ester of C6-C18 fatty acids derived from coconut oil, and mixtures thereof. Other hydrophobic substances also useful herein may be selected from monoglycerides, distilled monoglycerides, acetylated monoglycerides, diglycerides, triglycerides, and mixtures thereof. The hydrophobic substance used in the examples set forth herein for various zein coats was MCT oil, glycerine ester of C6-C18 fatty acids derived from coconut oil, manufactured by Karlshamns, of Columbus, Ohio, U.S.A., under the trade mark Captex 355.

Materials suitable for use in the enteric coating layer include enteric coating substances, with representative materials comprising methacrylic acid copolymer USP~NF Type A (also known as EudragitX L 100, Eudragit0 L 12.5, and Eudragit0 L 12.5P), methacrylic acid copolymer USP/NF
Type B (also known as Eudragit0 S 100, Eudragit S 12.5, and Eudragit~ S
12.5P), blends of methacrylic acid copolymer USP~NF Type A and methacrylic acid copolymer USP/NF Type B, methacrylic acid USP/NF Type C (also known as Eudragit~ L 30D and Eudragit8 L 100-55), hydroxypropyl mPthylcellulose, A

a1 225 9 2 -hydroxy propyl methyl cellulose phthalate, also known as HPMCP, HP50 or HP55, cellulose acetate phthalate, also known as C-A-P, cellulose acetate trimellitate, also known as C-A-T, blends of C-A-~ and C-A-P, polyvinyl acetate phthalate, also known as PVAP, and ethyl cellulose, also known as EC. Eudragit~ is an acrylic copolymer based on methacrylic acid and methyl methacrylate from Rohm Pharmac, of Germany, which has as a U.S.A. agent Rohm Tech, Inc. of Malden, Masschusetts. The foregoing examples are illustrative and not restrictive of suitable materials for inclusion in the delivery system of the invention, and the invention is considered to extend to unnamed equivalent materials within its scope. The enteric used herein was about a 3 to 1 weight/weight blend of Eudragit0 L 100 and Eudragi r S 100, respectively, which is believed to be the best mode of practicing the invention.
A plasticizer component for the enteric coat component may comprise for example triethyl citrate, acetyltriethyl citrate, tri-n-butyl citrate, acetyltri-n-butyl citrate, dibutyl phthalate, diethyl phthalate, dibutyl sebacate, glycerol triacetate, manufactured by Hoffmann La Roche under the trade name of Triacetin0, and acetylated monoglyceride, manufactured by Eastman Chemical Products under the trade mark of Myvacet~ 9-45. The plasticizer used herein was triethyl citrate.
An anti-tackiness agent for the enteric coat component comprises talci colloidal silca, and kaolin. The recommended level of anti-tackiness agents is preferably not greater than about 30% by weight of the total enteric compounds, in order to prevent the anti-tackiness agent from speeding disintegration of the enteric compound. The anti-tackiness agent used herein was Alpha-fil 500USP, talc manufactured by Cyprus Industrial Minerals Company, Englewood, Colorado, U.S.A.
The preparation of the active compound delivery system may be accomplished by a variety of coating techniques known in the art including fluid bed coating, coacervation, or a combination thereof, and the like, as disclosed in U.S. Pat. No. 4,384,004 to Cea et. al. Preferably, fluid bed coating with a rotor insert may be employed to form the initial core, and fluid bed coating with a Wurster column may be employed to apply the first, second, and third coatings. In the fluidized bed procedure, with rotor insert, for preparing a core containing an active substance as employed herein, the active substance or active substance in a matrix is charged as a powder onto a variable speed horizontal rotor disc in an apparatus that creates a upward air current or stream in which the particles have a rotary movement about an at least approximately vertical axis. The particles pass through a zone of finely atomized coating material which causes the passing particles to be coated. Additional solvents can be applied after the application of the coating material to better form particles of the desired size. Finally, rotor speed is increased and fluidization air volume and air temperature are also increased to both form the particles and obtain the desired level of dryness. The foregoing method and apparatus are known as a fluidized bed with rotor disc and are set forth in detail in the following U.S. patents; u.s. Pat. Nos. 4,323,312 and Re. 32,307.

.
In the fluidized bed with ~urster column procedure as applied herein for applying the various coatings, the cores produced with the fluidized bed with rotor insert described above, or other means, are suspended in an apparatus that creates a strong upward air current or stream in which the particles move. The stream passes through a zone of finely atomized coating material which causes the passing particles to be coated, after which the coated particles move upward through the Wurster column and then travel downward in a fluidized condition countercurrent to a flow of heated fluidized gas whereupon they are dried. The particles may reenter the upward stream for a further coating until the desired weight ratio of coating to active core has been obtained. The foregoing method and apparatus are known as the Wurster Process and are set forth in detail in the following U.S. Patents: U.S. Pat. Nos. 3,089,824; 3,117,027;

3,196,827 3,241,520 and 3,253,944.

The prolamine coating materials are prepared for use as a solution capable of being uniformly atomized. The solubility of zein requires a solvent with both polar and non-polar groups in the proper ratio. The proper ratio of polar .and non-polar groups can be obtained with single solvents or two or more solvent mixtures. Examples of suitable single solvents are acetic acid, lactic acid, propionic acid, and propylene glycol.
The aqueous alcohols are preferred as solvents in many applications.
Examples of suitable alcohol/water systems are methanol/water, ethanol/water, isopropanol/water, and n-butanol/water. To obtain complete a -WO g3/12772 2 1 2 2 S Y 2 PCT/US92/11160 solubility above the cloud point, the ratio of alcohol to water varies for each alcohol chosen and the mixed solvent final temperature. If desired, other ingredients such as plasticizers or hydrophobic substances may be added to improve the properties of the final coating. Suitable plasticizers include triethylene glycol, propylene glycol, oleic acid, lactic acid acetamide, ethylene glycol monooleate, glycerin, glycerol monostearate, dibutyl tartrate, and tricresol phosphate. Suitable hydrophobic substances include vegetable and animal fats, either unhydrogenated, hydrogenated, or partially hydrogenated, fatty acids, and glycerine esters of fatty acids, with representative materials comprising palm oil, palm kernel oil, soybean oil, rapeseed oil, rice bran oil, sunflower oil, safflower oil, coconut oil, castor oil, MCT oil, also known as glycerine ester of C6-C18 fatty acids derived from coconut oil, and mixtures thereof. Other hydrophobic substances also useful herein may be selected from monoglycerides, distilled mono and diglycerides, acetylated mono and diglycerides, diglycerides, triglycerides, and mixtures thereof. The plasticizer may be added in known effective amounts within the scope of the invention. In general, amounts of about 5% to about 25% by weight of the zein are suitable.
The enteric coating is preferably applied to a core or coated particle using the technique described above. Plasticizers and anti-tackiness agents may be added to improve the properties of the coating. The plasticizer component for the enteric coat used in the practice of the present invention includes triethyl citrate, acetyltriethyl citrate, tri-n-butyl citrate, acetyltri-n-butyl citrate, dibutyl phthalate, diethyl phthalate, dibutyl sebacate, glycerol triacetate, manufactured by Hoffmann La Roche under the trade name of Triacetin, and acetylated monoglyceride, manufactured by Eastman Chemical Products, Kingsport, Tennessee, U.S.A., under the trade name of Myvacet 9-45. The anti-tackiness agent for the enteric second coat component comprises talc, colloidal silca, and kaolin.
As used herein and in the claims when a coating component is stated as being as a percent of a particle, it should be understood that the coating component by itself is a weight percent of the particle including any prior coats. In the examples, all values of the weight percent of coating components were determined by analytical analysis. In addition to the stated weight percent of the coating component contained in the coat, the coat would also contain any specified plasticizer, hydrophobic substance, WO g3/12772 PCI`/US92/11160 15 2122~92 and anti-tackiness agent at the weight percent specified in the example.

~ Acetaminophen (APAP) is a well-known analgesic and antipyretic drug.
In the United States, it is available for non-prescription over-the-counter sale in conventional liquid, suppository, capsule, tablet and caplet dosage forms. The tablet and caplet dosage forms typically contain 325 mg acetaminophen as "regular strength" or 500 mg as "extra strength".
Normally, regular strength tablets or caplets are taken as one or two every four hours, and the extra strength tablet or caplets are taken as one or two every six hours. Ideally, it would be desirable to extend the dosing interval while maintaining the initial plasma concentrations achievable with conventional tablets or caplets. This would provide immediate and extended therapeutic analgesic or antipyretic effect and reduce the number of doses necessary, thereby making therapy more convenient.
Coated particles were prepared comprising acetaminophen from Mallinckrodt, Inc. St. Louis, Missouri, U.S.A., as a core material coated with zein (F 4000) and then further coated with an enteric substance to form dual coated particles. The acetaminophen cores were prepared by sieving granular acetaminophen to a particle size range of about 177-420 microns.
A solution of a coating material was prepared comprising zein (F 4000, Freeman Industries) plus MCT oil equaling 7.6X of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The acetaminophen cores were coated using a fluidized bed coating procedure in a 4~/6" fluid bed unit with a see-through main chamber, bottom spray, and Wurster column insert. The coating solution was applied to 500.0 grams of the acetaminophen cores at a rate that varied from about 6.7 to 7.5 grams/minute. The atomizing air pressure for the spray nozzle was about 172 kPa. The fluidizing inlet air temperature varied automatically between 45.0 C and 49.4C with a corresponding air discharge temperature of between 30.00C and 32.20C. The resultant coated particles were sieved into particles in the range of about 420-500 microns, and particles larger than 500 microns. The 420-500 micron size particles were designated APP3, and had a coating of 73% by weight zein thereon. The particles having a size greater than 500 microns were designated APP4, and had a coating of 86% by weight zein thereon.

The 420-500 micron particle portion of the batch (APP3) was then subjected to a second fluidized bed coating procedure. A solution o~ a second coating material was prepared comprising Eudragit~ L 100 and Eudragit~ S 100, in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total EudragitX, plus talc (Alpha-fil 500USP
from Cyprus Industrial Minerals Company) equaling 30/O by weight of the total Eudragit~, as a 12.0~ by weight solution of ethanol/acetone at a 88.7/11.4 weight/weight ratio. The solution of the second coating material was then applied to 350 grams of the zein coated acetaminophen particles at a rate of about 7.6 grams/minute to form dually coated particles. The atomizing air pressure for the spray nozzle was about 110 kPa. The fluidizing inlet air temperature automatically varied between 46.1OC and 48.90C with a corresponding air outlet temperature between 30.0OC and 33.9OC. The resultant dual coated particles were designated APP8 and comprised 73% by weight zein applied as a first coating, and 5% by weight Eudragit~ as a second coating. As a second product in this example, a coated particle was prepared comprising a core material of actaminophen, coated with a mixture of Eudragit~ RS 100 PM and Eudragit~ RL 100 PM, in a 4/1 weight/weight ratio, respectively, and then further coated with zein (F 4000) to form a reverse dual coated product of the present invention. The acetaminophen was prepared by sieving granular acetaminophen to a particle size range of about 177-420 microns. The resulting particles were then coated by fluidized bed coating procedure in a 4"/6" fluid bed unit with a see-through main chamber, bottom spray, and Wurster column insert. Accordingly, a mixture of EudragitX RS 100 PM and Eudragit~ RL 100 PM, in a 4/1 weight/weight ratio, respectively, plus triethyl cit~ate equaling 6% by weight of the total Eudragit~, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 15~ by weight of the total Eudragit~, was prepared as a 12.0% by weight solution of ethanol/acetone at a 94.4/5.6 weight/weight ratio, and applied to 800 grams of acetaminophen granular particles. The Eudragit~/triethyl citrate/talc solution was applied at a rate of about 6.8 to 7.0 grams/minute. The fluid atomizing air pressure was about`110 kPa.
The fluidizing inlet air temperature automatically varied between 46.1C and 47.2 C with a corresponding air outlet temperature between 30.6OC and 32.2-C. Samples were taken and are designated APP6, which has 22% by weight total Eudragit~ as Eudragit~ RS 100 PM/Eudragit~ RL 100 PM, 4/1 WO g3/12772 PCI`/US92/1~160 2122~92 weight/weight ratio, respectively; and APP7, which has 35% by weight total Eudragit~ as Eudragit RS 100 PM/Eudragit~ RL 100 PM, 4/1 weight/weight ratio, respectively.
~The sample represented by APP7 was then subjected to a second fluid bed coating. Zein (F 4000, Freeman Industries) was then applied to 415 -grams of the Eudragit0 coated acetaminophen to form reverse dually coated particles, with Eudragit~ next to the core as a first coat and zein on the outside as an exterior coating. Accordingly, zein (F 4000, Freeman Industries) plus MCT oil equaling 7.6% of the zein, was prepared as a 11.4%
by weight solution of ethanol/water at a 80/20 weight/weight ratio. It was then applied to 415.0 grams of Eudragit~ coated acetaminophen particles.
The zein/Mct oil solution was applied at a rate of about 6.5 grams/minute.
The fluidizing inlet air temperature varied automatically between 46.1C and 47.20C with a corresponding air discharge temperature of between 29.4OC and 33.9OC. The batch was not sieved. A sample was taken and designated APP9, 35% by weight Eudragit~ applied as a first coating, and then 75% by weight zein (F 4000) applied as a second coating.
The particles were subjected to dissolution tests in simulated gastric fluid (SGF) at pH 1.2 and simulated intestinal fluid (SIF) at pH 6.8. The SGF and SIF were prepared per USP specs (USP XXII - NF XVII, pp. 1788-1789, 1990). Selected samples were also tested in SGF at pH 5.0, in pH 1.2 and 5.0 acids, and in pH 6.8 buffer. The pH 1.2 and 5.0 acids were prepared the same way as the corresponding SGF's, except no enzymes were added.
Similarly, the pH 6.8 buffer was prepared the same way as the SIF, except no enzyme was added.
The dissolution test was conducted using apparatus 2 assembly, as specified by USP (USP XXII - NF XVII, pp. 1578-1579, 1990). The nominal capacity of the dissolution vessel was 1000 mL, ànd a paddle was used as the stirring element at a rotational speed of 50 rpm. The temperature was maintained at 37 + 0.5 oc using a circulating water bath.
The solution was sampled at 0.5, 1, and 2 hours for the test in SGF, and at 1, 3, and 6 hours for the test in SIF. A 10-mL aliquot was drawn at each sampling time through a 35-micron filter. The aliquot was then filtered again through a 0.45-micron filter for HPLC analysis to determine its drug content.

WO 93/12772 PCI`/US92/11160 An HPLC method was developed for the determination of acetaminophen in zein-coated powders and in dissolution samples from various media.
Zein-coated powders were dissolved in methanol for analysis. An acetaminophen USP reference standard was used for the quantitation. For zein-coated drug analysis, the standard was prepared in methanol. For dissolution sample analysis, the standard was prepared in water. Each sample or standard solution was then filtered through a 0.45-micron filter for HPLC analysis. A C-18 column connected with an UV detector at 244 nm was used for the HPLC analysis. The mobile phase was water/methanol (85/15, v/v) at a flow rate of 1.0 mL/min. The analysis time was 18-20 min/injection.
Dissolution results from the particles manufactured in Example 1 are set forth in Table 1.

WO 93tl2772 PCI~/US92~11160 2122~92 Dissolution Of Coated Acetaminophen In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release Of APAP
In SGF, pH 1.2 In SIF, pH 6.8 SampleX Drug 0.5 hr 1 hr 2 hr 1 hr 3 hr 6 hr APP3 55.9 29 47 79 24 44 67 APP4 52.0 -- -- -- 30 51 66 APP6 75.8 64 86 101 83 91 102 APP7 66.6 42 62 90 53 88 103 APP8 52.2 3 7 20 24 44 73 APP9 36.9 23 36 58 23 43 65 APP21* 100.0 97 97 98 98 101 102 *uncoated particles of Acetaminophen The particles from sample APP9, which had a dual coating with the inner coat being an enteric compound and the exterior coating being a prolamine (zein), did exhibit sustained release, as compared to APP21 which is uncoated, in simulated intestinal fluid. Photomicrographs made of various structures from other of the Examples herein, after the zein coated particles were subjected to dissolution media showed that the zein did not totally disintegrate or dissolve, but remained in place throughout the dissolution. The photomicrographs in combination with the dissolution results from various particles, which will be discussed in later examples, indicate that the respective locations of the prolamine and enteric layers may be varied in order to facilitate the desired dosing protocol for a given active substance to a particular patient population.

Dual coated particles were prepared using the same procedure as set forth in Example 1. In this instance, acetaminophen cores were prepared by W O 93/12772 P(~r/uss2/lll6o sieving granular acetaminophen to a particle size range of about 177-420 microns. The first coating material was again zein (F 4000, Freeman Industries) and MCT oil, prepared as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio, and applied to 800 grams of acetaminophen at a rate of about 6.2 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature initially varied automatically between 45.0OC and 47.8OC with a corresponding air discharge temperature of between 27.8C and 32.2OC.
Because of high humidity, the fluidizing inlet air temperature was raised to between 73.3OC and 73.9OC, with a corresponding air outlet temperature of between 36.1C and 45.0OC, in an attempt to control agglomeration. This increase in temperature appeared to be effective in reducing some of the agglomeration. Finally, in an attempt to further control agglomeration, the fluidizing inlet air temperature was again raised to between 74.4OC and 78.3 C, with a corresponding air outlet temperature of between 39.4OC and 43.9OC, that appeared to reduce agglomeration further. Preferably, the inlet air relative humidity is controlled to be less than 40%, if such a control is available on the equipment being used. The resultant particles were sieved and were designated: APP10, which comprised a coating of 22% by weight zein, with the particles having sizes of less than 500 microns;
APP11, which comprised a coating of 24X by weight zein, with the particles having sizes of less than 500 microns; APP12, which comprised a coating of 31% by weight zein, with the particles having sizes of less than 500 microns; APP15, which comprised a coating of 34% by weight zein, with the particles having sizes of about 500-590 microns; APP16, which comprised a coating of 33% by weight zein with the particles having sizes in the range of about 500-590 microns; and APP17, which comprised a coating of 64% by weight zein, with the particles having sizes in the range of about 500-590 microns.
The particles represented by the sample designated APP12, were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit~ L 100 and Eudragit S 100 in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit, plus talc (Alpha-fil 500USP
from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit~, was prepared as a 12.0% by weight solution of ethanol/acetone at WO 93/12772 PCI`/US92~11160 a 88.7/11.4 weight/weight ratio. The second coating material was then applied to 553 grams of the zein coated acetaminophen cores at a rate of about 6.4 grams/minute to form dually coated particles. The atomizing air pressure for the spray nozzle was about 110 kPa. The fluidizing inlet air temperature automatically varied between 46.1C and 49.4C with a corresponding air outlet temperature between 29.4OC and 30.60C. At various times throughout the coating procedure samples were taken and sieved, and were designated: APP13, which comprised 31% by weight zein applied as a first coat followed by 18Yo by weight Eudragit as a second coat, with the particles having sizes of less than 500 microns; APP14, which comprises 31% by weight zein applied as a first coat followed by 307O by weight Eudragit~ as a second coat, with the particles having sizes of less than 500 microns; APP18, which designates 31% by weight zein as a first coat, followed by 39% by weight Eudragit~ as a second coat, with the particles having sizes in the range of about 500-590 microns; and APP19, which comprises 31% by weight zein applied as a first coat, followed by 56%
by weight Eudragit as a second coat, with the particles having sizes in the - range of about 500-590 microns.
The particles produced in this example were subjected to dissolution testing as described in Example 1, and the results are set forth in Table 2.
Selected samples were subjected to sequential dissolution. The sample was first in simulated gastric fluid (SGF) for 1 hour, and then in simulated intestinal fluid (SIF) for an additional 6 hours. Sampling time was at 0.5 and 1 hour in SGF and at 1, 3, and 6 hours in SIF.
Two methods were used for the sequential dissolution. In one method, a 40-mesh basket (USP apparatus 1 assembly) was used as the stirring element, in which the powder sample was placed. In the other method a paddle (USP apparatus 2 assembly) was used as the stirring element.
In the basket method, the dissolution medium was changed from SGF to SIF during the transition. In the paddle method, less SGF was initially used which was neutralized to pH 6.8 at the end of 1 hour. A pre-warmed and more concentrated SIF solution was then added to make a SIF solution at the right concentration for the subsequent dissolution test. The paddle method was modified from an USP procedure for Delayed-release drugs (USP XXII - NF
XVII, pp. 1580-1581, 1990).

WO 93/12772 PCI`/US92/11160 2122~92 Dissolution Of Coated Acetaminophen In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release Of Acetaminophen In SGF, pH 1.2 In SIF, pH 6.8 SamDle % Drug 0.5hr lhr 2hr lhr 3hr 6hr APP10 80.7 74 86 89 70 84 88 APP11 79.8 66 83 87 50 67 76 APP12 75.1 64 78 85 51 65 73 -- 76 79 79*
7279**
APP13 59.5 9 11 17 33 67 78 4 6 -- 36 58 71*
APP14 52.4 8 11 15 39 73 85 9 12 -- 43 60 69*
APP15 73.4 40 68 85 33 63 85 APP16 73.9 46 72 97 41 68 84 APP17 59.3 27 46 74 25 44 68 APP18 47.9 4 6 8 32 53 77 APP19 41.3 3 4 6 24 56 76 APP21 100.0 97 97 98 98101102***

* sequential dissolution procedure ** dissolution performed in pH 8.4 buffer *** uncoated particles of Acetaminophen The dissolution results for particles from samples APP3, APP10, APP12 and APP21 in simulated gastric fluid are presented in graphic form in Figure 1. These samples were selected as being representative of the samples manufactured in Examples 1 and 2. The data for APP21 clearly demonstrates that uncoated particles of acetaminophen (APAP) are dissolved in gastric WO 93/12772 PCI`/US92/11160 fluid very quickly. When the APAP cores were coated with a single layer of zein (APP3, APP10, APP12) the rate of dissolution of the APAP was slowed, but a majority of the APAP was released from the core within two hours.
APP10, APP12 and APP3 had coating of 22%, 31% and 73%, respectively, zein as a weight percentage of the APAP core. Heavier coatings of zein slowed the release of the core material more than lighter coatings. However, it is an objective of the present invention to minimize the release of the active substance in the stomach, and coating the active substance with zein alone does not meet this objective. This objective is very significant when the active substance is one, such as a ~-lactim antibiotic, which preferably has minimal contact with the acidic environment of the stomach. In consideration of the fact that the absorptive capacity of the stomach is only about lX of the absorptive capacity of the intestinal tract, release of the active substance in the intestinal tract is very desirable if a rapid release of the active substance in the stomach is not indicated by the dosing protocol.
Figure 2 is a graphic presentation of the dissolution rates of particles from samples APP3, APP10, APP12 and APP21 in simulated intestinal fluid. (These dissolutions were done with fresh particles, not those used in the dissolution results in simulated gastric fluid shown in Figure 1.) Once again, the uncoated APAP particles (APP21) were dissolved fairly rapidly. The particles coated with zein released the APAP from the cores much slower in a sustained release fashion. The particles having heavier coatings of zein on the cores had a more uniform sustained rate of release (APP3 as compared to APP10 and APP12). It is clear from this data that the quantity of the zein coating must be varied in order to achieve the desired rate of sustained release of an active substance in the intestinal tract.
It is interesting to note from Table 2 that particles from sample APP12 were also subjected to a sequential dissolution procedurej which confirmed that a majority of an active substance coated only with zein is released in the gastric fluid.
Figure 3, when compared to Figure 1, shows a significant reduction in the rate of dissolution of APAP when the cores are coated first with a layer of zein and then with a layer of an enteric substance. The APP8 particles are APP3 particles having an exterior layer of an enteric substance thereon, and the APP13 particles are APP12 particles having an exterior layer of an WO 93/12772 PCI~/US92/1 1 160 2l22s92 enteric substance thereon. As compared with the uncoated APAP particles (APP21) the enteric coated particles released a minimal amount of the active substance in the simulated gastric fluid.
Figure 4, when compared to Figure 2, shows that the enteric coatings on particles from the same samples as shown in Figure 3 did not significantly impair the sustained release characteristics in simulated intestinal fluid which are shown in Figure 2.
It is to be noted from Table 2 that dual coated particles from sample APP13 were subjected to a sequential dissolution test which shows that dual coated particles of the type made in Example 2 do in fact restrict release of an active substance in simulated gastric fluid while exhibiting the desired sustained release characteristic in the simulated intestinal fluid.
It is believed that the dissolution rates of the APAP were affected by the fact that the APAP cores were elongated and needle-like in shape rather than spherical. This core shape resulted in thinner coatings at the "ends" of the cores, which affected dissolution results.
It may be fairly concluded from the data presented thus far that a composite structure for delivery of an active substance for sustained release in the intestinal tract in accordance with the present invention may comprise a core containing an active substance, said core being coated with a prolamine and said prolamine being coated with a layer of an enteric substance. It is understood that the amount of the coating materials is dependent upon the desired dosing protocol for the active substance.

Dual coated particles of the type prepared in Example 2 were further coated to form triple coated particles. In this instance, acetaminophen cores were prepared by sieving granular acetaminophen to particles having sizes in the range of about 125-250 microns. The first coating material was again zein (F 4000, Freeman Industries) and MCT oil; however, prepared as a 12% by weight solution of ethanol/water at a ratio of 87/13 weight/weight, and applied to 756 grams of acetaminophen at a rate of about 5.8 grams/minute. The atomizing air pressure for the spray nozzle was about 138 kPa. The fluidizing inlet air temperature varied automatically between 68.3 C and 76.70C with a corresponding air discharge temperature of between 42.20C and 47.2C. A second batch of coated particles was prepared WO 93/12772 PCl'/US92/11160 212259~

duplicating this procedure, and the two batches were intimately mixed in the fluid coating apparatus, after which additional zein coating was applied.
At separate times during the coating procedure samples were taken, but not sieved and were designated: APP22, having a coating of 32% by weight zein applied; and APP23, having a coating of 46X by weight zein.
The particles designated APP23, were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragi r S 100 in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit, as a 12.0%
by weight solution of ethanol/acetone at a 88.7/11.4 weight/weight ratio.
The solution of the second coating material was then applied to 846 grams of the zein coated acetaminophen particles at a rate that varied from about 5.5 to 6.0 grams/minute to form dually coated particles. The atomizing air pressure for the spray nozzle was about 110 kPa. The fluidizing inlet air temperature was automatically varied between 47.80C and 52.2 C with a corresponding air outlet temperature between 31.7C and 35.6 C. A sample was taken and sieved and was designated APP24, which comprised 46% by weight zein applied as a first coat, followed by 28% by weight Eudragit~ as a second coat, with the particles having sizes of less than 500 microns.
The particles designated APP24 were split in two batches. One of those batches was subjected to a third fluidized bed coating procedure. A
solution of a third coating material was prepared comprising zein (F 4000, Freeman IndustriesJ and MCT oil; as a 12X by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution of the third coating material was applied to 374 grams of the APP24 particles at a rate of about 5.4 to 6.5 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature varied automatically between 75.6OC and 78.3OC with a corresponding air discharge temperature of between 41.1C and 47.2OC. At various times throughout the coating procedure samples were taken, sieved, and designated as follows:
APP25, which comprises 46% by weight zein (F 4000) applied as a first coat followed by 28% by weight Eudragit as a second coat followed by 23% by weight zein (F 4000) as a third coat, with the particles having sizes of less than 500 microns; APP26, which comprised 46% by weight zein (F 4000) WO 93/12772 2 1 2 2 5 9 2 PCI`/US92/11160 _ applied as a first coat followed by 28% by weight Eudragit~ as a second coat followed by 7% zein (F 4000) applied as a third coat, with the particles having sizes of less than 297 microns; and APP27, which designates 46% by weight zein (F 4000) as a first coat followed by 28% by weight Eudragit~ as a second coat followed by 41% zein (F 4000) as a third coat, with the particles having sizes in the range of about 297-420 microns.
The other batch of particles from the sample designated APP24, was then subjected to a third fluidized bed coating procedure. A solution of a third coating material was prepared comprising a different zein (F 4000LE, Freeman Industries) and MCT oil, prepared as a 12% by weight solution of ethanol/water at a ratio of 87/13 weight/weight, and applied to 338 grams of the APP24 particles at a rate of about 5.2 to 5.7 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature varied automatically between 75.60C and 78.3OC with a corresponding air discharge temperature of between 41.1C and 45.6OC. At various times throughout the coating procedure samples were taken, sieved, and designated as follows: APP30, which comprised 46% by weight zein (F 4000) applied as a first coat followed by 28% by weight Eudragit as a second coat followed by 18% by weight zein (F 4000LE) as a third coat, with the particles having sizes of less than 420 microns; and APP32, which comprised 46% by weight zein (F 4000) applied as a first coat followed by 28X by weight Eudragit~ as a second coat followed by 28Yo zein (F 4000 LE) applied as a third coat, with the particles having sizes of less than 420 microns.
The particles manufactured in this example were then subjected to dissolution testing as described in Example 1. Selected particle samples were subjected to sequential dissolution testing as described in Example 2.
The results of these dissolution tests are set forth in Table 3.

WO 93/12772 PCr/US92/11160 ~_ 27 Dissolution Of Coated Acetaminophen In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release Of Acetaminophen In SGF, pH 1.2 In SIF, pH 6.8 SampleX Drug 0.5 hr 1 hr 2 hr 1 hr 3 hr 6 hr APP21 100.0 97 97 98 98 101 102 APP22 74.1 65 90 97 42 78 90 APP23 66.7 63 87 98 26 51 65 62 88 -- 96 94 94*
APP24 47.8 6 10 18 47 88 104 6 11 -- 36 54 63*
APP25 38.2 11 16 23 26 60 80 APP26 44.4 15 23 34 33 67 82 APP27 32.9 11 15 24 17 35 55 11 15 -- 41 78 93*
APP30 40.0 7 10 18 12 33 55 APP32 36.5 11 16 26 21 41 57 9 14 -- 32 50 66*

*sequential dissolution procedure WO 93/12772 PCI`/US92/11160 Figures 5 and 6 are graphs showing the dissolution of triple coated particles manufactured in Example 3 in simulated gastric fluid and simulated intestinal fluid, respectively. In both of these graphs APP21 represents the dissolution of uncoated particles of acetaminophen (APAP). Particles from samples APP25 and APP32 have a first coating layer of zein on an APAP
core, then a layer of an enteric substance over the first zein layer, and then a second coating of zein as an exterior coating on the particles. The desirability of an exterior coating of zein will be better explained in Example 11 wherein such particles are placed in liquid suspensions. At this point it can be briefly stated that an exterior coating of zein will protect the enteric coating when the particles are placed in a liquid medium having a pH of greater than about 5Ø When Figure 5 is compared to Figure 3 it is clear that an exterior coating of zein does not significantly impair the property of the enteric coating for minimizing the release of an active substance from the core in simulated gastric fluid. Furthermore, when Figure 6 is compared to Figure 4, it is clear that the exterior coating of zein does not significantly impair the desired sustained release of the active substance in simulated intestinal fluid, although the rate of release may be slowed.
Figure 7 is a graphic representation of the dissolution of an active substance (APAP) when triple coated particles (APP27 and APP32) are subjected to the sequential dissolution procedure as compared to particles having only a single coating of zein on the core (APP23). It can be concluded from this data that the triple coated particles exhibit a minimal rate of release in the simulated gastric fluid followed by the desired sustained rate of release in simulated intestinal fluid. The outer coating on the APP32 particles is a zein having a lower ash content than that in the outer coating of the APP27 particles. This data indicates that a slower rate of sustained release is attained when a zein having a lower ash content is used in coating the particles.

In this example, single coated particles were prepared comprising a core material of acetaminophen, coated with zein (F 4000), varying the level of the hydrophobic substance, MCT oil, and also substituting plasticizers at different levels for the MCT oil. In this example, acetaminophen cores were prepared by sieving granular acetaminophen to a particle size range of about 125-297 microns.
A solution of a coating was prepared comprising zein (F 4000, Freeman Industries) and MCT oil, which is a hydrophobic plasticizer, at 7.6% by weight of the zein; as a 25% by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution of the coating material was applied to 750 grams of the acetaminophen cores at a rate between 5.3- 11.4 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 58.30C and 65.6C with a corresponding air discharge temperature of between 32.80C and 37.8C. A sample was taken and sieved, and was designated APP36, which comprised a coating of 37Z by weight zein (F 4000) with the particles having sizes of less than 297 microns.
A solution of a coating material was prepared comprising zein (F 4000, Freeman Industries) and MCT oil at 5% by weight of the zein, as a 24.6% by weight solution of ethanol/water at a ratio of 87/13 weight/weight, and was applied to 750 grams of acetaminophen cores at a rate between 6.0-6. 3 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 56.1-C and 61.1oC with a corresponding air discharge temperature of between 35.0-C and 38.3C. A sample was taken and sieved, and was designated APP35, which comprised a coating of 44% by weight zein (F 4000) with the particles having sizes of less than 297 microns. A solution of a coating material was prepared comprising zein (F 4000, Freeman Industries) and MCT oil at 20% by weight of the zein; prepared as a 25.0% by weight solution of ethanol/water at a ratio of 87/13 weight/weight; and was applied to 750 grams of acetaminophen cores at a rate between 6.0-6.1 grams/minute.
The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 55.00C and 61.7C with a corresponding air discharge temperature of between 31.1OC and 38.3-C. Less plugging of the fluid nozzle occurred than at the lower levels of MCT oil; however, particle agglomeration appeared equivalent, again likely due to the 25.0% by weight solution being too high. A sample was taken and sieved, and was designated APP37, which comprised a coating of 41%

W O 93tl2772 PC~r/US92/11160 2122~92 30 by weight zein (F 4000) with the particles having sizes of less than 297 microns.
A fourth batch of particles was prepared using a combination of Myvacet~ 9-45 and Myverol P-06, which are amphoteric substances, in a 4/1 weight/weight ratio at 7.6% by weight of the zein, as a replacement for the MCT oil. Both Myvacet0 9-45 and Myverol P-06 are products manufactured by Eastman Chemicals. A solution of a coating material was prepared comprising zein (F 4000, Freeman Industries) and a combination of Myvacet 9-45 and Myverol P-06 in a 4/1 weight/weight ratio at 7.6% by weight of the zein, as a 25.0% by weight solution of methanol/acetone/water at a ratio of 82/14/4 weight/weight/weight, and was applied to 750 grams of acetaminophen at a rate between 6.6-6.8 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 52.2OC and 56.1C with a corresponding air discharge temperature of between 31.1C and 35.6OC. A sample was taken and sieved, and was designated APP38, which comprised a coating of 45% by weight zein (F 4000) with the particles having sizes of less than 297 microns.
A fifth batch of particles was prepared using a combination of Myvacet 9-45 and Myverol P-06, in a 4/1 weight/weight ratio at 20.0% by weight of the zein, as a replacement for the MCT oil. A solution of a coating material was prepared comprising zein (F 4000, Freeman Industries) and a combination of Myvacet~ 9-45 and Myverol P-06 in a 4/1 weight/weight ratio at 20% by weight of the zein, as a 24.2% by weight solution of methanol/acetone/water at a ratio of 82/14/4 weight/weight/weight, and was applied to 750 grams of acetaminophen at a rate of between 6.2-8.9 grams/minute. The atomizing air pressure for the spray nozzle was about 83 kPa. The fluidizing inlet air temperature varied automatically between 53.5OC and 60.6OC with a corresponding air discharge temperature of between 31.7C and 35.6OC. A sample was taken and sieved, and was designated APP39, which comprised coating of 46% by weight zein (F 4000) with the particles having sizes of less than 297 microns.
The particles produced in this example were subjected to dissolution testing as described in Example 1. The results of these dissolution tests are set forth in Table 4.

WO 93/~2772 PCr/US92/11160 ~ 31 2 1 2 2 5 9 2 Dissolution Of Coated Acetaminophen In Simulated Gastric - (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release Of APAP
Plasticizer In SGF, pH 1.2 In SIF, pH6.8 Sample % Drug Hydrophobic Amphoteric 0.5 hr 1 hr 2 hr 1 hr 3 hr 6 hr APP21* 100.0 100.0 - 97 97 98 98 101 102 APP35 68.4 5.0 - 72 95 99 70 98 103 APP36 71.8 7.6 - 69 92 98 72 91 93 APP37 66.9 20.0 - 61 89100 60 87 96 APP38 67.5 - 7.6% 71 97 99 66 94 100 APP39 64.2 - 20% 68 96102 65 95 103 *uncoated APAP

In all of these samples an attempt was made to have a substantially constant level of zein applied as 37-46% of the weight of the core, with the hydrophobic substance and levels varying, or substituting an amphoteric plasticizer for the hydrophobic substance. A first conclusion is that the rate of release initially decreased in both SGf and SIF with increasing levels of MCT oil, however this effect is modest and becomes almost non-apparent in both SGF and SIF over a period of time, and that the differences are not significant. There appears to be no significant difference if plasticizer is substituted for MCT oil.
To prevent pure zein from cracking, the zein needs to be mixed with a plasticizer or hydrophobic, and these can interchangeably be used.
However, if there is a critical active substance for which leakage needs to be restricted as much as possible the MCT oil or the hydrophobic substance is preferred.

W O 93/12772 pc~r/us92/ll16o In this example, triple coated particles, of the type manufactured in Example 3, were manufactured. In this example the effect of zein with two different levels of ash content was evaluated in various coating layers, "Zein LE" desingates zein from Freeman Industries having an ash content equal to or less than 0.1%, and "Zein" is also from Freeman Industries and has an ash content of up to 2%. In this instance, a granular acetaminophen with a particle size range of about 125-250 microns was used for the cores.
No further sieving was performed. The first coating material was again zein (F 4000, Freeman Industries) and MCT oil; however, prepared as a 20% by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The first coating solution was applied to 800 grams of acetaminophen at a rate of about 3.2-4.9 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 58.3OC and 61.7C with a corresponding air discharge temperature of between 35.0C and 43.3 C. Minimum agglomeration and no nozzle plugging occurred. A sample taken, but not sieved, was designated APP40, which comprised a single coating of 23% by weight zein (F 4000) applied to the core.
Particles from the sample designated APP40, were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit~ L 100 and Eudragit S 100 in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit~, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30X by weight of the total Eudragit, as a 10.7X by weight solution of ethanol/acetone/water at a 87.0/10.0/3.0 weight/weight/weight ratio. This second coating material was then applied to 500 grams of the APP40 particles at a rate of about 5.0 grams/minute to form dually coated particles. The atomizing air pressure for the spray nozzle was about 110 kPa. The fluidizing inlet air temperature automatically varied between 40.6OC and 55.6OC with a corresponding air outlet temperature between 30.0OC and 33.9OC. Minimum agglomeration and no nozzle plugging occurred. A sample was taken, but not sieved, and was designated APP41, which comprised 23% by weight zein (F 4000) applied as a first coat followed by 19% by weight Eudragit~ as a second coat.
Particles from the sample designated APP41 were then subjected to a third fluidized bed coating procedure. A solution of a third coating material was prepared comprising zein (F 4000LE, Freeman Industries) and MCT
oil at 7.6% by weight of the zein; as a 20% by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution of the third coating material was applied to 500 grams of the APP41 particles at a rate of about 5.3 to 5.4 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa to 138 kPa. The fluidizing inlet air temperature varied automatically between 53.5OC and 63.3OC with a corresponding air discharge temperature of between 34.40C and 39.4C. A sample was taken and sieved and was designated APP42, which comprised 23X by weight zein (F 4000) applied as a first coat followed by 19% by weight Eudragit as a second coat followed by 23% by weight zein (F 4000LE) as a third coat, with the particles having a size of less than 297 microns.
A second batch of particles was then prepared, again using the granular acetaminophen comprising particles having sizes in the range of about 125-250 microns as cores. A solution of a first coating material was prepared comprising zein (F 4000LE, Freeman Industries) and MCT oil at 7.6%
by weight of the zein; as a 20X by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution of the first coating material was applied to 800 grams of acetaminophen cores at a rate of about 5.3 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa. The fluidizing inlet air temperature varied automatically between 59.4OC and 61.1C with a corresponding air discharge temperature of between 35.00C and 42.8OC. Minimum agglomeration and no nozzle plugging occurred.
A sample was taken, but not sieved, and was designated APP44, comprising a single coating of 20% by weight zein (F 4000LE).
Particles from the sample designated APP44, were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit S 100 in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit, as a 10.7% by weight solution of ethanol/acetone/water at a 87.0/10.0/3.0 weight/weight/weight ratio. The solution of the second coating material was WO 93/12772 PCr/US92/11160 then applied to 490 grams of the APP44 particles to form dually coated particles at a rate of about 4.9-5.0 grams/minute. The atomizing air pressure for the spray nozzle was about 110 kPa. The fluidizing inlet air temperature automatically varied between 45.60C and 46.1C with a corresponding air outlet temperature between 28.9OC and 33.90C. Minimum agglomeration and no nozzle plugging occurred. A sample were taken but not sieved and was designated APP45, which comprised 20% by weight zein (F
4000LE) applied as a first coat, followed by 16% by weight Eudragit as a second coat.
Particles represented by the sample designated APP45 were subjected to a third fluidized bed coating procedure. A solution of a third coating material was prepared comprising zein (F 4000LE, Freeman Industries) and MCT
oil at 7.6% by weight of the zein; as a 20% by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution of the third coating material was applied to 500 grams of the APP45 particles at a rate of about 5.3 to 5.5 grams/minute. The atomizing air pressure for the spray nozzle was about 97 kPa to 138 kPa. The fluidizing inlet air temperature varied automatically between 53.5OC and 63.3OC with a corresponding air discharge temperature of between 34.4OC and 39.4OC. A sample was taken and sieved and was designated APP46, which comprised 20% by weight zein (F
4000LE) applied as a first coat followed by 16% by weight Eudragit~ as a second coat followed by 23% by weight zein (F 4000LE) as a third coat, with the particles having sizes of less than 297 microns.
The particles produced in this example were subjected to dissolution testing as described in Example 1. Selected particle samples were subjected to sequential dissolution testing as described in Example 2. The results of the dissolution tests are set forth in Table 5.

WO 93/12772 PCI`/US92/11160 Dissolution Of Coated Acetaminophen In Simulated Gastric (pH 1.2J And Intestinal Fluids (pH 6.8) Cumulative % Release Of Acetaminophen In SGF, pH 1.2 In SIF, pH 6.8 Sample % Drug 0.5 hr1 hr 2 hr 1 hr 3 hr 6 hr 1st coat 3rd coat APP21 100. 97 97 98 98 101102 - -APP40 80.2 82 94 99 70 98 103 23%R
APP41 62.8 17 22 29 87 103103 23%R
APP42 50.4 14 19 -- 44 63 71* 23%R 23%R
APP44 82.0 72 91 98 72 98 101 20%LE
APP45 66.7 15 17 22 88 101101 20%LE
APP46 53.7 17 22 -- 49 69 74* 20%LE 23%LE

*sequential dissolution procedure It may be concluded that at these levels of zein coatings the use of R and LE zein does not significanlty effect dissolution results. However, at higher levels of zein, (see samples APP27 and APP32 from Example 3) but in a sequential dissolution for APP27 and APP32 there is a difference because the zein having a lower ash content (LE) slowed the dissolution rate. The ash content of zein and the amount of coating can be varied to achieve the desired dosage protocol for any particluar active substance.

WO 93/12772 PCl`/US92/11160 In this example, single coated particles were prepared comprising a core material of acetaminophen, coated with zein from different manufacturers: zein from Freeman Industries (F 4000LE) comprising at least 90% protein; zein from Corn Products (lot 1952) comprising at least 90YO
protein; and zein from Nutrilite (lot 011-09 comprising about 87% protein).
Both Corn Products of Pekin, Illinois, U.S.A., and Nutrilite of Buena Park, California, U.S.A., no longer manufacture zein. Granular acetaminophen was used with a particle size range of about 125-250 microns.
A first batch of particles was prepared using zein from Freeman Industries (F 4000LE). A solution of a coating material was prepared comprising zein (F 4000LE, Freeman Industries) and MCT oil at 7.6X by weight of the zein; prepared as a 20% by weight solution of ethanol/water at a ratio of 87/13 weight/weight. The solution was applied to 800 grams of acetaminophen cores at a rate between 5.0-5.8 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature varied automatically between 53.50C and 68.9OC with a corresponding air discharge temperature of between 29.4OC and 46.1C. At various times during the coating procedure samples were taken and were designated: APP48, which comprised coating of 23% by weight zein (F 4000LE);
APP49, which comprised a coating of 31X by weight zein (F 4000LE); and APP50, which comprised a coating of 46% by weight zein (F 4000LE).
A second batch of particles was prepared using zein from Corn Products. A solution of a coating material was prepared comprising a 20.0%
by weight solution of ethanol/water at a ratio of 87/13 weight/weight, and applied to 1200 grams of acetaminophen. The solution was applied to the acetaminophen cores at a rate of about 6.1 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature varied automatically between 50.0OC and 55.6OC with a corresponding air discharge temperature of between 26.1C and 34.4C.
Samples were taken and were designated: APP54, which comprised a coating of 22X by weight zein (Corn Products); APP55, which comprised a coating of 30%
by weight zein (Corn Products); and APP56, which comprised a coating of 43%
by weight zein (Corn Products).
A third batch of particles was prepared using zein from Nutrilite (lot 011-09). A solution of a coating was prepared comprising a 20.0% by weight WO 93/12772 PCI`/US92/11160 solution of ethanol/water at a ratio of 87/13 weight/weight. The solution was applied to 1200 grams of acetaminophen cores at a rate of about 6.1-6.2 grams/minute. The atomizing air pressure for the spray nozzle was about 124 kPa. The fluidizing inlet air temperature varied automatically between 53.50C and 58.30C with a corresponding air discharge temperature of between 29.40C and 35.00C. Samples were taken and were designated: APP57, which comprised a coating of 24% by weight zein (Nutrilite); APP58, which comprised a coating of 34% by weight zein (Nutrilite); and APP59, which comprised a coating of 53% by weight zein (Nutrilite).
The particles manufactured in this example were subjected to dissolution testing as described in Example 1. The results of the dissolution tests are set forth in Table 6.

WO 93/12772 PCI`/US92/11160 Dissolution Of Coated Acetaminophen In Simulated Intestinal Fluid (pH 6.8) Sample % Drug 1 hr 3 hr 6 hr APP21 100.98 101 102 APP48 80.1 70 98 102 APP49 74.9 47 82 94 APP50 66.7 57 84 100 APP54 81.0 63 99 103 APP55 75.5 57 81 90 APP56 68.5 51 79 90 APP57 79.8 90 101 104 APP58 73.5 85 98 101 APP59 63.8 79 95 100 Depending upon the source of zein, which may be extracted by different processes, all zein exhibits varying amounts of effect on the rate of sustained release. The th;ckness and type of coating may be varied to achieve the desired dosage protocol for a particular active substance.

The present invention has special utility as a delivery system for ~-lactam antibiotics, such as amoxicillin, which should be protected from the acidic environment of the stomach. As utilized hereinafter the term N~_ lactam antibioticsN shall mean compounds having a beta-lactam ring as a central structure, i.e., the structure ~ N

WO 93/12772 PCI~/US92/11160 which thereafter may be substituted at various positions on the ring and/or fused with other ring systems which may themselves be substituted or unsubstituted. Some examples of well-known ~-lactam antibiotics include penicillins, cephalosporins, monocyclic ~-lactams, e.g. azthreonam, thienamycin and its derivatives, and the clavulanic acid derivatives as well as the pharmaceutically acceptable salts of the above-mentioned compounds.
In this example, dual coated particles were prepared comprising a corematerial of amoxicillin trihydrate from Interchem Corporation, Paramus, New Jersey, U.S.A., granulated with zein (F 4000, from Freeman Industries) using a Glatt GPCG 3 and top spray. A solution of a granulating material was prepared comprising zein, MCT oil and amoxicillin trihydrate, with MCT
oil equaling 7.6% of the zein, and amoxicillin equaling 80% of the zein, as a 27.1% by weight solution of ethanol/water at a 80/20 weight/weight ratio.
The granulating material was then applied to 750.0 grams of amoxicillin trihydrate particles at a rate of about 40.0 grams/minute. The fluidizing inlet air temperature varied automatically between 60.0OC and 61.1C with a corresponding product temperature of between 25.0OC and 48.3OC. The particles were sieved to be in a size range of about 100-200 microns. A
sample was taken and designated AMX2, amoxicillin granulated with 60% by weight zein. The particles of sample AMX2 were then coated with zein (F
4000) and then further coated to form a dual coated product of the present invention. A solution of a first coating material was prepared comprising zein (F 4000, Freeman Industries) plus MCT oil equaling 7.6% of the zein, as a 11.4X by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution of the first coating material was then applied to 750.0 grams of the AMX2 cores at a rate that varied from about 5.5 to 8.4 grams/minute using a fluidized bed coating procedure in a Glatt GPCG 3 with a 17.8 cm Wurster column insert, and bottom spray. The fluidizing inlet air temperature varied automatically between 45.00C and 46.1C with a corresponding particle temperature of between 21.1C and 35.0OC. Samples were taken and were designated: AMX3, which comprised AMX2 as the core with a single coat of 47% by weight Zein; and AMX4, which comprised AMX2 as the core with a single coat of 51% by weight zein.
The AMX4 particles were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit S 100, in a 3/1 WO 93/12772 PCI`/US92/11160 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit~, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit, as a 12.0% by weight solution of ethanol/acetone at a 88.7/11.3 weight/weight ratio. The solution of the second coating material was then applied to 750 grams of the AMX4 particles at a rate of about 11.6-11.8 grams/minute to form dually coated particles. The fluidizing inlet air temperature automatically varied between 46.1C and 47.8OC with a corresponding air outlet temperature between 40.0OC and 43.3OC. Samples were designated: AMX5, which comprised AMX2 as the core, with 51% by weight zein applied as a first coating and 39%
by weight Eudragit as a second coating; and AMX6, which comprised AMX2 as the core, with 51% by weight zein applied as a first coating and 45% by weight Eudragit~ as a second coating.
The particles manufactured in this example were subjected to dissolution testing as described in Example 1. Selected particle samples were subjected to sequential dissolution testing as described in Example 2.
A USP HPLC method (USP XXII - NF XVII, supplement 1, pp.
2088-2089,1989) was adapted for the determination of amoxicillin in both zein-coated powders and dissolution samples from various media. The column was C-18. The mobile phase was a pH 5.0 potassium phosphate buffer containing 4% acetonitrile, at a flow rate of 1.5 mL/min. The detector was UV at 230 nm. A pure amoxicillin trihydrate (AMX1), which was calibrated with an USP reference standard, was used as the working standard. For dissolution sample analysis, the standard was dissolved in the moile phase.
For zein-coated drug analysis, both the sample and the standard were dissolved by mixing with 5 mL methanol for 2-5 minutes and then diluting with the mobile phase to the desired level.
The results of the dissolution tests are set forth in Table 7.

W O 93/12772 2 1 2 2 S 9 2 P(~r/us92/lll6o -Dissolution Of Coated Amoxicillin In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) .

Cumulative % Release Of Amoxicillin In SGF, pH 1.2 In SIF, pH 6.8 Sample% Drug 0.5 hr 1 hr 2 hr 1 hr 3 hr 6 hr AMX2 60.7 -- -- -- 86 85 89 AMX3 40.2 -- -- -- 36 66 79 42 60 66**
AMX4 39.1 -- -- -- 50 66 77 46 43 -- 80 79 79*
56 62 63**
AMX5 25.1 18 18 21 46 61 71 21 -- 49 60 66*
AMX6 23.6 12 20 25 37 62 75 18 27 -- 69 75 78*
AMX23 100.0 53 45 36 83 91 93***

* sequential dissolution procedure ** dissolution performed in pH 8.4 buffer *** uncoated particles The dissolution results for the particles made in this example showed that sustained release in simulated intestinal fluid is not achieved merely by granulating an active substance with zein, even at zein levels as high as 65X of the total weight (see AMX2 in Table 7). However, when zein was used as a coating material on this same granulated core, then sustained release in simulated intestinal fluid was achieved.

W O 93/12772 PC~r/US92/11160 In this example, dual coated particles were prepared comprising a core material of amoxicillin trihydrate rotor granulated with zein (F 4000, from Freeman Industries) using a Glatt GPCG 5 with rotor insert. A solution of a granulating material was prepared comprising zein plus MCT oil, with MCT
oil equaling 7.6% of the zein, was prepared as a 6.0% by weight solution of ethanol/water at a 80/20 weight/weight ratio. With the rotor turning at 500RPM, the solution containing 250 grams of zein, was applied to 2.5 kilograms of micronized amoxicillin trihydrate particles, at a rate that varied from about 50.0-65.0 grams/minute. The atomizing air pressure for the spray nozzle varied from 179 kPa to 221 kPa. The fluidizing inlet air temperature varied automatically while spraying between 30.0C and 35.0C
and a corresponding product temperature while spraying between 16.1C and 20.0C. Following completion of spraying of the zein solution, 700 grams of distilled water/ethanol at a 57/43 weight/weight ratio respectively, was sprayed on the batch at a rate of about 85.0 grams/minute.
The atomizing air pressure for the spray nozzle was about 179 kPa.
After completion of the water/ethanol spraying, drying air was increased to between 50.0OC and 75.0OC with a corresponding product temperature while drying between 26.1-C and 32.20C. The granules were sieved to sizes of: (a) less than 180 microns; (b) greater than 250 microns; and (c) between 180-250 microns.
A solution consisting of the amoxicillin granules greater than 250 microns was prepared as a 11.7% by weight slurry of distilled water/ethanol at a 83/17 weight/weight ratio, respectively. Using a Glatt GPCG 5 with rotor insert, the slurry was applied to 1.703 kilograms of the sieved amoxicillin granules less than 180 microns. With the rotor turning at 500 RPM, the amoxicillin slurry was applied at a rate that varied from about 50.0-62.0 grams/minute. The atomizing air pressure for the spray nozzle varied from 179 kPa to 221 kPa. A total of 240 grams of ethanol/distilled water at a 80/20 weight/weight ratio was again applied prior to drying at a rate of about 62.0 grams/minute. The atomizing air pressure for the spray nozzle was about 200 kPa. Drying was terminated when product temperature reached 32.20C to 33.9C. The fluidizing inlet air temperature while spraying was about 30.0 C, while drying between 40.0C and 75.00C with a corresponding granule temperature while spraying between 17.8C and 20.00C, W O 93/12772 Pc~r/uss2/lll6o ~ 432122~92 and while drying between 26.1C and 32.2 C. The granules were sieved to sizes of: (a) less than 125 microns; and (b) greater than 250 microns; and (c) between 125-250 microns. The 180-250 micron size granules from both the first and second charges were intimately mixed and sampled. This sample is designated AMX31, amoxicillin rotor granules with 11% by weight zein.
The core particles of sample AMX31 were then coated with zein (F 4000) and then further coated to form a dual coated product of the present invention. A solution of a first coating material was prepared comprising zein (F 4000, Freeman IndustriesL plus MCT oil equaling 7.6X of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The particles were coated by fluidized bed coating procedure in a Glatt GPCG 1 with a 15.2 cm Wurster column insert, and bottom spray. The solution of the first coating material was applied to 529.0 grams of the AMX31 amoxicillin cores at a rate of about 12.0 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 47.80C and 52.80C with a corresponding product temperature of between 27.8OC and 36.70C. The particles were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit S 100, in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15X by weight of the total Eudragit, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit, as a 12.0X by weight solution of ethanol/acetone at a 88.7/11.3 weight/weight ratio. The solution of the second coating material was applied to 620 grams of the single coated particles to form dually coated particles at a rate of about 12.0 grams/minute. The atomizing air pressure for the spray nozzle was about 303 kPa. The fluidizing inlet air temperature automatically varied between 47.8OC and 48.90C with a corresponding product temperature between 36.1C
and 41.1C. Samples were taken and designated as follows: AMX32, which comprised AMX31 as the core, with 45X by weight zein applied as a first coating and 22X by weight Eudragit as a second coating; and AMX33, which comprised AMX31 as the core, with 45X by weight zein applied as a first - coating and 52% by weight Eudragit as a second coating.
The particles were subjected to dissolution as described in Example 1. The dissolution results are presented in Table 8.

WO 93/12772 PCI'/US92/11160 Dissolution Of Coated Amoxicillin In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release of Amoxicillin In SGF, pH 1.2 In SIF, pH 6.8 Sample % Drug 0.5 hr 1 hr 2 hr 1 hr 3 hr 6 hr AMX23 100.0 53 45 36 83 91 93**
AMX31 89.5 40 33 25 91 88 86*
AMX32 45.5 3 4 9 24 53 77 AMX33 34.5 3 2 3 20 42 66 * agglomerated core without coating ** uncoated, unagglomerated core The dissolution results for particles from samples AMX23 (uncoated, unagglomerated amoxicillin) and AMX32 (agglomerated amoxicillin core, first coat zein and exterior coat of an Eudragit~ compound) are presented in Figures 8 and 9. In simulated gastric fluid (Figure 8) the uncoated AMX23 had a rapid release, but the release of the active substance from the double coated AMX32 was minimal. This confirms the dissolution results for dual coated APAP cores shown in Figure 3.
The dissolution results for AMX23 and AMX32 in simulated intestinal fluid are presented in Figure 9. Once again, as already demonstrated in Figure 4 for APAP, the active substance was released in sustained manner from the double coated core, but quite rapidly when the active substance was not coated.

WO 93/12772 P(~r/US92/11160 ~~ 4~1 2 2 5 9 2 In this example, triple coated particles were prepared comprising a core material of amoxicillin trihydrate rotor granulated with zein (F 4000, from Freeman Industries) using a Glatt GPCG S with rotor insert. A solution of a granulating material was prepared comprising zein plus MCT oil, with MCT oil equaling 7.6% of the zein, as a 6.0Yo by weight solution of ethanol/water at a 80/20 weight/weight ratio. To this granulating solution was added 445 grams of amoxicillin trihydrate. With the rotor turning at 500 RPM, the slurry containing 250 grams of zein and 445 grams of amoxicillin was applied to 2~055 grams of amoxicillin trihydrate micronized particles at a rate that varied from about 52.0-63.0 grams/minute. The atomizing air pressure for the spray nozzle varied from about 166 kPa to 186 kPa. Following the zein/amoxicillin slurry, 200 grams of distilled water/ethanol at a 83/17 weight/weight ratio was then applied at a rate of about 63.0 grams/minute. The fluidizing inlet air temperature varied automatically while spraying between 38.9C and 40.0C~ and during drying about 50.00C with a corresponding granule temperature while spraying between 15.0C and 17.2C~ and while drying about 32.8C. The granules were sieved to less than 180 microns, greater than 250 microns, and between 180-250 microns. A slurry consisting of 386 grams of the amoxicillin granules greater than 250 microns was prepared as a 11.7X by weight slurry of distilled water/ethanol at a 83/17 weight/weight ratio. Using a Glatt 6PCG
5 with rotor insert, the slurry was applied to 1~295 grams of the sieved amoxicillin granules less than 180 microns. With the rotor turning at 500 RPM, the amoxicillin slurry was applied at a rate of about 52.0 grams/minute. The atomizing air pressure for the spray nozzle was about 166 kPa. A total of 200 grams of ethanol/distilled water at a 80/20 weight/weight ratio was applied prior to drying at a rate of about 62.0 grams/minute. The atomizing air pressure for the spray nozzle was about 166 kPa. Drying was terminated when product temperature reached 32.20C. The fluidizing inlet air temperature while spraying was about 40.0OC, while drying about 50.0C with a corresponding granule temperature while spraying between 17.8C and 20.0C~ and while drying about 32.20C. The granules were sieved to sizes of: (a) less than 180 microns; (b) greater than 250 microns;
and (c) between 180-250 microns. A slurry consisting of 100 grams of the amoxicillin granules greater than 250 microns was prepared as a 11.7% by WO 93/12772 PCr/US92/11160 weight slurry of distilled water/ethanol at a 83/17 weight/weight ratio.
Using a Glatt GPCG 5 with rotor insert, the slurry was applied to 1300 grams of the sieved amoxicillin granules having sizes of less than 180 microns. With the rotor turning at 500 RPM, the amoxicillin slurry was applied at a rate of about 52.0 grams/minute. The atomizing air pressure for the spray nozzle was about 166 kPa. A total of 200 grams of ethanol/distilled water at a 80/20 weight/weight ratio was again applied prior to drying at a rate of about 62.0 grams/minute. The atomizing a;r pressure for the spray nozzle was about 166 kPa. Drying was terminated when product temperature reached 32.2OC. The fluidizing inlet air temperature while spray;ng was about 40.0OC, and while drying about 50.0OC with a corresponding granule temperature while spraying between 16.7C and 20.00C, and while drying about 32.20C. The granules were sieved to sizes of: (a) less than 180 microns; and (b) greater than 250 microns; and (c) between 180-250 microns.
The 180-250 micron size granules the first, second, and third charges were intimately mixed and sampled. This sample was designated AMX34-1, amoxicillin rotor granules with 17% by weight zein.
Particles from sample AMX34-1 were then coated with zein (F 4000), then further coated to form a dual coated product of the present invention, and finally further coated to form a triple coated product of the present invention. The particles were coated by fluidized bed coating procedure in a Glatt GPCG 1 with a 15.2 cm Wurster column insert, and bottom spray. A
solution of a first coating material was prepared comprising zein (F 4000, Freeman Industries) plus MCT oil equaling 7.8% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 600 grams of rotor granules represented by sample AMX34-1 at a rate of about 15.0 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 46.7OC and 138F with a corresponding product temperature of between 22.20C and 37.2C.
Samples were taken and designated as follows: AMX35B, comprising AMX34-1 as the core, with 17% by weight zein applied as a first coating; and AMX36B, comprising AMX34-1 as the core, with 30% by weight zein applied as the first coating. The particles represented by sample AMX36B were then subjected to a second fluidized bed coating procedure.

WO 93/12772 PCl`/US92/11160 A solution of a second coating mater~al was prepared comprising a mixture of Eudragit L 100 and Eudragit~ S 100, in a 3/1 weight/weight ratio, plus triethyl citrate equaling 15% by weight of the total Eudragit2, as a 12.0% by weight solution of ethanol/acetone/water at a 84.2/11.3/4.5 weight/weight/weight ratio. The solution of the second coating material was applied to 700 grams of the single coated AMX36B particles to form dually coated particles at a rate of about 14-16.0 grams/minute. The atomizing air pressure for the spray nozzle was about 303 kPa. The fluidizing inlet air temperature automatically varied between 47.8C and 61.1C with a corresponding product temperature between 31.1C and 38.9DC. Samples were taken and designated as follows: AMX37B, which comprised AMX34-1 as the core, with 30~, by weight zein applied as a first coating and 8% by weight Eudragit~ as a second coating; and AMX38B, which comprised AMX34-1 as the core, with 30% by weight zein applied as a first coating and 16% by weight Eudragit~ as a second coating. A solution of a third coating material was prepared comprising zein (F 4000LE, Freeman Industries) plus MCT oil equaling 7.6% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 655 grams of the dual coated particles of AMX38B at a rate between about 14.0-16.0 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 47.2OC and 60.0C with a corresponding product temperature of between 26.1C and 41.1C.
Samples were taken and designated as follows: AMX39, comprising AMX34-1 as the core, with 30% by weight zein (F 4000) applied as a first coating, 16% by weight Eudragit~ as a second coating, and 74% zein (F
4000LE) as a third coating; and AMX40, comprising AMX34-1 as the core, with 307O by weight zein (F 4000) applied as a first coating, 16% by weight Eudragit~ as a second coating, and 76% zein (F 4000LE) as a third coating.
A second batch of triple coated particles was then produced using the following procedure. A second batch of amoxicillin rotor particles were produced using the same conditions as used in making the sample designated AMX34-1, but with a different ratio of zein to amoxicillin. This sample was designated AMX34, amoxicillin rotor granules with 30% by weight zein.
Particles from sample AMX34 were then coated with zein (F 4000), then further coated to form a dual coated product of the present invention, and WO 93/12772 PCr/US92/11160 finally further coated to form a triple coated product of the present invention. The particles were coated by fluidized bed coating procedure in a Glatt GPCG 1 with a 15.2 cm Wurster column insert, and bottom spray. A
solution of a first coating material was prepared comprising zein (F 4000, Freeman Industries) plus MCT oil equaling 7.6% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 750 grams of rotor granules represented by sample AMX34 at a rate of about 15.0 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 46.70C and 58.9OC with a corresponding product temperature of between 17.2C and 35.C.
The particles were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit~ S 100, in a 3/1 weight/weight ratio, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit2, plus triethyl citrate equaling 15% by weight of the total Eudragit~, as a 12.0X by weight solution of ethanol/acetone/water at a 84.2/11.3/4.5 weight/weight/weight ratio. The solution of the second coating material was applied to 750 grams of the single coated particles to form dually coated particles at a rate of about 10.0-16.0 grams/minute. The atomizing air pressure for the spray nozzle was about 303 kPa. The fluidizing inlet air temperature automatically varied between 48.9C and 60.0OC with a corresponding product temperature between 28.9C and 37.20C. Samples were taken and designated as follows: AMX42, which comprised AMX34 as the core, with 10% by weight zein applied as a first coating and 14X by weight Eudragit~ as a second coating; and AMX43, which comprised AMX34 as the core, with 10% by weight zein applied as a first coating and 23X by weight Eudragit~ as a second coating. A solution of a third coating material was prepared comprising zein (F 4000LE, Freeman Industries) plus MCT oil equaling 7.6% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 560 grams of the dual coated particles of AMX43 at a rate between about 13-21 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 47.8C and 53.9 C with a corresponding product temperature of between 25.0C and 37.80C. Samples were taken and designated as follows: AMX44, comprising AMX34 as the core, with 10Z by weight zein (F 4000) applied as a first coating, 23% by weight Eudragit~ as a second coating, and 75% zein (F 4000LE) as a third coating;
and AMX45, comprising AM%34 as the core, with lOYo by weight zein (F 4000) applied as a first coating, 23% by weight Eudragit as a second coating, and 95% zein (F 4000LE) as a third coating.
A third batch of triple coated particles was then produced using the following procedure. The same batch of amoxicillin rotor particles designated by sample AMX34, amoxicillin rotor granules with 30% by weight zein, was used to produce this triple coated product. Particles from sample AMX34 were coated with zein (F 4000), then further coated to form a dual coated product of the present invention, and finally further coated to form a triple coated product of the present invention. The particles were coated by fluidized bed coating procedure in a Glatt GPCG 1 with a 15.2 cm Wurster column insert, and bottom spray. A solution of a first coating material was prepared comprising zein (F 4000, Freeman Industries) plus MCT
oil equaling 7.6% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 325 grams of rotor granules represented by sample AMX34 at a rate of about 15.0 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 48.90C and 60.0OC with a corresponding product temperature of between 17.2C and 33.9OC. A sample was taken and designated AMX46, which comprised AMX34 as the core, with 80% by weight zein applied as a first coating. The particles were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit~ S 100, in a 3/1 weight/weight ratio, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit~, plus triethyl citrate equaling 15% by weight of the total Eudragit~, as a 12.0%
by weight solution of ethanol/acetone/water at a 84.2/11.3/4.5 weight/weight/weight ratio. The solution of the second coating material was applied to 275 grams of the single coated particles of AMX46 to form dually coated particles at a rate of about 10.0 grams/minute. The atomizing air pressure for the spray nozzle was about 303 kPa. The fluidizing inlet air temperature automatically varied between 48.9OC and 60.00C with a WO 93/12772 PCI~/US92/11160 212259~

corresponding product temperature between 32.2C and 42.2C. Samples were taken and designated as follows: AMX47, which comprised AMX34 as the core, with 80% by weight zein applied as a first coating and 12% by weight Eudragit as a second coating; and AMX48, which comprised AMX34 as the core, with 80% by weight zein applied as a first coating and 20% by weight Eudragit as a second coating. A solution of a third coating material was prepared comprising zein (F 4000LE, Freeman Industries) plus MCT oil equaling 7.7% of the zein, as an 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 277 grams of the dual coated particles at a rate of about 14 grams/minute. The atomizing air pressure for the spray nozzle was about 283 kPa. The fluidizing inlet air temperature varied automatically between 47.8OC and 58.9OC with a corresponding product temperature of between 25.00C
and 37.80C. Samples were taken and designated as follows: AMX49, comprising AMX34 as the core, with 80% by weight zein (F 4000) applied as a first coating, 2 Wo by weight Eudragit as a second coating, and 7 Wo zein (F
4000LE) as a third coating; and AMX50, comprising AMX34 as the core, with 80% by weight zein (F 4000) applied as a first coating, 2 WO by weight Eudragit as a second coating, and 78% zein (F 4000LE) as a third coating.
The particles produced in this example were subjected to dissolution in the manner set forth in Example 1. Some particles were subjected to sequential dissolution in the manner set forth in Example 2. The results are presented in Table 9.

WO 93/12772 PCI`/US92/11160 Dissolution Of Coated Amoxicillin In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release Of Amoxicillin In SGF, pH 1.2 In SIF, pH 6.8 Sample% Drug 0.5 hr 1 hr 2 hr 1 hr 3 hr6 hr AMX34 75.6 61 48 36 103 108 108 AMX35B 71.4 31 22 5 102 100 93 AMX36B 64.1 27 6 4 76 87 87 AMX37B 57.4 51 46 32 81 82 84 AMX38B 52.3 14 4 4 79 86 86 AMX3g 29.2 21 28 -- 44 51 51*
AMX40 28.8 16 23 -- 36 45 45*
AMX42 56.6 47 43 36 104 111 109 AMX44 28.4 11 15 -- 29 40 48*
AMX45 25.7 9 15 -- 36 57 66*
AMX48 31.6 33 40 -- 55 62 67*
AMX49 18.1 6 10 -- 16 20 26*
AMX50 17.2 4 7 -- 12 15 23*

*sequential dissolution procedure The sequential dissolution results for triple coated particles from sample AMX45 are presented in a graph in Figure 10. The desired sustained release of the active substance in simulated intestinal fluid (SIF) was achieved, while there was minimal release of the active substance in simulated gastric fluid (SGF). This confirms the sequential dissolution results for triple coated APAP which are presented in Figure 7.
It was further concluded from the dissolution results presented in Table 9 that both of the zein coating layers in a triple coated particle contribute to the rate of release in simulated intestinal fluid. (Compare the dissolution results for AMX39, AMX40, AMX44, AMX49 and AMX50.) WO 93/12772 P(~r/US92/11160 In this example, dual coated particles were prepared comprising a core material of amoxicillin trihydrate rotor granulated with zein (F 4000, from Freeman Industries) using a Glatt GPCG 5 with rotor insert. A solution of a granulating material was prepared comprising zein plus MCT oil, with MCT
oil equaling 7.6Yo of the zein, as a 6.0% by weight solution of ethanol/water at a 80/20 weight/weight ratio. With the rotor turning at 300 RPM, the slurry containing 150 grams of zein was applied to 2,500 grams of micronized amoxicillin trihydrate particles at a rate about 51 grams/minute. The atomizing air pressure for the spray nozzle was about 221 kPa. Following the spraying of the zein solution, the unit was shut down without drying the batch. The Glatt GPCG 5 filter was discharged into a sheet of plastic sheeting. This 442 grams of filter product was then slurried into 1,625 grams of ethanol/distilled water at a 60/40 weight/weight ratio. This slurry was then applied at a rate of about 51.0 grams/minute. Distilled water was then sprayed at about 51 grams/minute until the rotor torque reached 52.0 Newton-Meters. At this point the rotor RPM was increased from 300 to 500 RPM and drying was started. The fluidizing inlet air temperature while spraying the zein solution and the filter particle slurry was about 30.0-C, and during drying between about 50.00C and 75.00C with a corresponding granule temperature while spraying between 9.4C and 21.1C, and while drying about 35.C. The granules were sieved to less than 100 microns, greater than 125 microns, and between 100-125 microns. The 100-125 micron size granules were sampled. This sample is designated AMX62, amoxicillin rotor granules with 7% by weight zein.
Particles from sample AMX62 were then coated with zein (F 4000LE), then further coated to form a dual coated product of the present invention.
~he particles were coated by a fluidized bed coating procedure in a Glatt GPCG 1 with a 15.2 cm Wurster column insert, and bottom spray. A solution of a first coating material was prepared comprising zein (F 4000LE, Freeman Industries) plus MCT oil equaling 7.6% of the zein, as a 11.4% by weight solution of ethanol/water at a 80/20 weight/weight ratio. The solution containing zein was applied to 500 grams of rotor granules represented by sample AMX62 at a rate of about 9-10 grams/minute. The atomizing air pressure for the spray nozzle was about 324 kPa. The fluidizing inlet air temperature varied automatically between 5~.7OC and 52.8C with a WO 93~12772 PCl/US92/11160 corresponding product temperature of between 32.2OC and 37.8 C. A sample was taken and designated AMX63, which comprised AMX62 as the core, with 21%
by weight zein (F 4000LE) applied as a first coating. The particles represented by sample AMX63 were then subjected to a second fluidized bed coating procedure. A solution of a second coating material was prepared comprising a mixture of Eudragit L 100 and Eudragit0 S 100, in a 3/1 weight/weight ratio, plus talc (Alpha-fil 500USP from Cyprus Industrial Minerals Company) equaling 30% by weight of the total Eudragit, plus triethyl citrate equaling 15% by weight of the total Eudragit, as a 12.0%
by weight solution of ethanol/acetone/water at a 84.2/11.3/4.5 weight/weight/ weight ratio. The solution of the second coating material was applied to 469 grams of the single coated particles to form dually coated particles at a rate of about 8-10.0 grams/minute. The atomizing air pressure for the spray nozzle was about 303 kPa. The fluidizing inlet air temperature automatically controlled to 50.0OC, with a corresponding product temperature between 33.9OC and 36.1C. Samples were taken and designated:
AMX64, which comprised AMX62 as the core, with 21X by weight zein (F 4000LE) applied as a first coating and 29X by weight Eudragit as a second coating;
and AMX65, which comprised AMX62 as the core, with 21% by weight zein (F
4000LE) applied as a first coating and 67% by weight Eudragit as a second coating.

Dissolution Of Coated Amoxicillin In Simulated Gastric (pH 1.2) And Intestinal Fluids (pH 6.8) Cumulative % Release of Amoxicillin In SGF, pH 1.2 In SIF, pH 6.8 Sample% Drug 0.5 hr 1 hr2 hr 1 hr 3 hr 6 hr AMX63 76.4 54 46 -- 47 48 50 AMX64 53.6 3 6 -- 14 22 35 AMX65 38.8 2 2 -- 6 14 29 WO 93~12772 PCI`/US92/11160 Only zein having a very low ash content by weight of less than 0.1%
(zein F 4000LE from Freeman Industries) was used in the particles made in this example. Even when used sparingly, the lower ash content zein had a positive effect in retarding the release of the active substance from the core. Lower ash content zein may be used when the size of particles due to coating thicknesses becomes a critical factor in a particular application of the invention.

Amoxicillin trihydrate was coated with different combinations and concentrations of zein (regular and LE grades) and Eudragit (L100 plus S100 in a ratio of 3:1 weight/weight) as shown in Table 11. The particles designated AMXl are uncoated particles of amoxicillin. A description of the manufacture of the AMX65 particles is set forth in Example 10; for the AMX44 and AMX49 particles in Example 9.
The purpose of this example is to demonstrate that particles in accordance with the invention may be disposed in a liquid medium to provide an alternative system for delivery of an active substance for release in the intestinal tract. As used herein and in the claims, a Hliquid medium" is understood to be an oil based liquid, aqueous based liquid, or a liquid that has a base which is a combination of water and oil. As used herein and in the claims, a "liquid" is understood to mean a state of matter in which the molecules are relatively free to change their positions with respect to each other but are restricted by cohesive forces to maintain a relatively fixed volume.

W O 93/12772 PC~r/US92/11160 Composition of Microencapsulated Amoxiciltin Products Tested in Suspension Stability Study Sample ID Percent Coating Composition and Concentrations Code AmoxicillinFirst Coat Second Coat Third Coat Zein LE Eudragit AMX 65 38.8 21 67 ~Zein Regular Eudragit Zein LE
AMX 44 28.4 10 23 75 AMX 49 18.1 80 20 70 Samples of particles having each of the structures from Table 11 were suspended at a concentration of 250 mg amoxicillin per 5 mL in sucrose-based syrups adjusted to different pH's, (the formulations of these syrups are set forth in Table 12), were stored at 30OC and shaken daily. Samples were removed for testing at 1 hour, 1 day, 7 days and 14 days. Encapsulated drug and undissolved particulate matter were removed from the suspensions by filtration through a 35 micron filter. The clarified syrups were diluted with mobile phase of pH 5.0 potassium phosphate buffer containing 4%
acetonitrile and filtered again through a 0.45 micron filter prior to analysis by high pressure liquid chromatography (HPLC). The method of quantitating the amount of amoxicillin released in the syrup solutions was adapted from the USP method for amoxicillin described on pages 2088-2089 of Supplement 1 in USP XXII - NF XVII.

Wo 93/12772 P(~r/uss2/lll6o Composition of Syrups for Microencapsulated Amoxicillin Suspension Stability Study IngredientSyrup A Syrup B Syrup C

pH 5.0 + 0.1 pH 6.5 + 0.1pH 8.5 + 0.1 Sucrose 89.21 gm 89.21 gm 89.21 gm Potassium Sorbate 0.595 gm 0.595 gm 0.595 gm Xanthan Gum 0.222 gm 0.222 gm 0.222 gm Citric Acid 1.033 gm 0.101 gm Sodium Citrate 3.332 gm 6.319 gm ----Sodium Bicarbonate ---- ---- 1.478 gm Sodium Carbonate ---- ---- 0.1 gm Water 100 mL 100 mL 100 mL

WO 93/12772 PCI~/US92/11160 2122~92 Table 13 shows that very little amoxicillin was released from samples AMX4, AMX49 and AMX65 during the test period. Each of these samples had at least one coating layer of zein encapsulating the core. Sample AMX1 was uncoated amoxicllin. This was true regardless of the pH of the syrup. It is believed that at least some of the "release" at one hour was actually fines and uncoated particles from the coating process. The syrup formulation was designed to minimize the solubility of amoxicillin so one would not expect a high rate of release (see results of the uncoated amoxicillin, AMX1). The slow rate of release in the syrup, however, is thought to be largely due to the 67 percent overcoat of Eudragit in AMX65, and the 75 and 70 percent zein LE outer coats in AMX44 and AMX49, respectively.

WO 93/12772 PC~r/US92/11160 2l2 2S9 2 58 Effect of Coating Composition and Syrup pH on the Release of Amoxicillin Microencapsulated with Two Kinds of Zein and Eudragit~

Sample Percent Percent Amoxicillin Released in Syrup Code Amoxicillin (Storage Time) 1 Hr 1 Day 7 Days 14 Days Syrup at pH 5.0 AMX 1 100 7.4 9.3 11.3 9.0 AMX 65 38.8 0.8 1.1 1.2 0.5 AMX 44 28.4 1.5 1.7 2.5 1.7 AMX 49 18.1 0.3 0.3 1.2 1.1 Syrup at pH 6.5 AMX 1 100 8.6 9.6 14.6 10.9 AMX 65 38.8 1.8 1.6 2.4 1.4 AMX 44 28.4 1.9 2.0 2.9 1.8 AMX 49 18.1 0.2 1.0 2.1 1.3 Syrup at pH 8.5 AMX 1 100 19.5 2.8 9.6 5.7 AMX 65 38.8 3.0 0.3 1.5 2.3 AMX 44 28.4 2.0 0.1 0.7 0.5 AMX 49 18.1 0.3 0.1 1.3 0.7 WO 93/12772 PCI~/US92~1160 The concept that stability in an aqueous suspension is primarily controlled by the exterior coating layer is supported by studies performed on acetaminophen (APAP). Acetaminophen was coated with various combinations of zein and Eudragit0. Table 14 shows the formulations of the syrups in which the various paricles were suspended. The composition of these syrups was similar to those used for the Amoxicillin suspensions (compare Tables 12 and 14). A description of the manufacture of the APP27 particles is set forth above in Example 3; for APP32 particles in Example 3; for APP42 particles in Example 5; and for APP46 particles in Example 5. The particles identified in the tables as "APAP" are uncoated acetaminophen. APP27 and 32 were suspended at 250 mg per 5 mL in a syrup at pH 8.1 and APP42 and APP46 were each suspended at the same concentration in syrups at both pH 5.0 and 8.1 respectively.
For analysis of samples in sucrose-based syrups, the samples were diluted 50-400 folds with water and then filtered through a 0.45-micron filter for HPLC analysis. The dilution was necessary for proper UV
detection and to minimize the effect of matrix on column performance. The methanol content of the aqueous mobile phase was adjusted between 2 and 15%, pending the performance of the C-18 column used.
Tables 15 and 16 show that the particles were not coated well enough to prevent leakage of the drug into the syrup. They do show that the highest concentrations of regular or LE grade zein in the outer coat have the lowest amount of acetaminophen released. The underlying coats may contribute to the relative stability of APP27 and APP32, but it appears that stability must be controlled predominately by the concentration of the outer coat when one compares the release of drug and the concentration of the first and second coating layers of APP42 and APP46 with AMX44 (Tables 16 and 13).

WO 93/12772 PCr/US92/11160 -2 ~2 2 5 9 2 TABLE 14 Composition of Syrups per 150 mL for Microencapsulated Acetaminophen Suspension Stability Studies Ingredient Syrup A Syrup B

pH 5.0 pH 8.1 Sucrose 96.727 gm 96.727 gm Potassium Sorbate 0.645 gm 0.645 gm Xanthan Gum 0.242 gm 0.242 gm Silica Gel 0.322 gm 0.322 gm Sodium Bicarbonate 1.612 gm 1.612 gm Flavoring 0.451 gm 0.451 gm Deionized Water 97.8 mL 100.0 mL
Concentrated HCl 2.2 mL --Approximate Volume 150.0 mL 150.0 mL

W O 93~12772 PC~rJUS92/11160 61 2 1 2 2 ~ 9 2 Effect of Different Coating Concentrations of Zein and Eudragit~ on the Stability of Microencapsulated APAP in a Syrup at pH 8.1 Percent APAP Released in Syrup Sample Percent (Storage TimeJ
Code APAP 1 Hr. 5 Hr. 1 Day 7 Days14 Days28 Days APP 27 32.9 1.4 1.8 2.7 5.4 5.6 8.1 APP 32 36.5 1.6 2.7 4.8 8.1 10.0 12.0 APAP 99. 6 20.0 ND ND ND ND 23.0 Syrup O O ND ND ND ND O

ND = Not Determined WO 93/12772 PCr/US92/11160 zi22s92 62 Effect of Different Coating Concentrations of Zein and Eudragit~ on the Stability of Microencapsulated APAP in Syrups at pH 5.0 and 8.1 Sample Percent pH Percent APAP Released in Syrup Code APAP (Storage Time) 1 Hr 1 Day 7 Days 14 Days APP 42 50.4 5.0 4.0 11.315.0 15.0 8.1 3.2 12.319.8 20.5 APP 46 53.7 5.0 7.2 13.417.5 16.2 8.1 3.5 16.316.5 28.0 APAP 99.6 5.0 20.2 31.926.5 24.9 8.1 7.7 27.529.8 28.0 This example shows that a suspension containing particles having an active substance in a core coated with at least one layer of a prolamine and one layer of an enteric substance has been reduced to practice, with minimal leakage of the active substance into the suspension medium.

Claims (33)

The embodiments of the invention in which an exclusive property or privilege is claimed are defined as follows:

1. A system for delivery of an active substance for release in the intestinal tract comprising a particle having a core containing an active substance, said core being encapsulated by a first layer of a coating material containing an enteric substance in an amount from about 10% to 70% of the total weight of the core, and said first coating layer being encapsulated by an exterior layer of a coating material containing a prolamine and at least one of a plasticizer and a hydrophobic substance, said prolamine being in an amount of about 20% to 100% by weight of the sum of the weights of the core and the first coating layer.

2. A system according to claim 1, wherein the enteric compound comprises at least one material selected from the group consisting of acids and esters of methacrylic copolymers.

3. A system according to claim 1, wherein the prolamine is zein.

4. A system according to claim 2, wherein the prolamine is zein.

5. A system according to claim 3, wherein the zein has an ash content of not greater than about 2%
by weight.

6. A system according to claim 2, wherein the zein has an ash content of not greater than about 2%
by weight.

7. A system according to claim 1, 2, 3, 4, 5 or 6, wherein the active substance is selected from the group consisting of analgesics, antibiotics, antidepressants, antivirals, antibodies, immuno-modulators, oncolytics, immunogens, hormones, vaccines, enzymes, nutrients and dietary fiber.

8. A system according to claim 1, 2, 3, 4, 5 or 6, wherein the active substance is a .beta.-lactam antibiotic.

9. A system according to claim 1, 2, 3, 4, 5 or 6, wherein said exterior layer contains MCT oil.

10. A system according to claim 1, 2, 3, 4, 5 or 6, wherein said amount of said enteric substance is 20% to 40%, by weight, of the total weight of the core; and said prolamine is in an amount of about 40%
to 60%, by weight, of the sum of the weights of the core and the first coating.

11. A system according to claim 10, wherein said exterior layer contains MCT oil.

12. A system for delivery of an active substance for release in the intestinal tract comprising a particle having a core containing an active substance, said core being encapsulated by a first layer of a coating material containing a coating material containing a prolamine and at least one of a plasticizer and a hydrophobic substance, said prolamine being in an amount of about 10% to 70% of the total weight of the core, and said first coating being encapsulated by a second coating layer containing an enteric compound at about 10% to 70% of the sum of the weights of the core and the first coating layer.

13. A system according to claim 12, wherein the particle is in the form of a tablet.

14. A system according to claim 12, wherein the prolamine is zein.

15. A system according to claim 13, wherein the prolamine is zein.

16. A system according to claim 12, 13, 14 or 15, wherein said first layer contains MCT oil.

17. A system according to claim 12, 13, 14 or 15, wherein said first layer contains about 20% to 50%, by weight, of said prolamine, based on the total weight of the core; and said second layer contains about 20% to 60%, by weight, of said enteric compound, based on the sum of the core and the first coating.

18. A system according to claim 12, 13, 14 or 15, wherein the active substance is selected from the group consisting of analgesics, antibiotics, antidepressants, antivirals, antibodies, immuno-modulators, oncolytics, immunogens, hormones, vaccines, enzymes, nutrients and dietary fiber.

19. A system according to claim 12, 13, 14 or 15, wherein the active substance is a .beta.-lactam antibiotic.

20. A system according to claim 17, wherein said first layer contains MCT oil.

21. A system according to claim 18, wherein said first layer contains MCT oil.

22. A system according to claim 19, wherein said first layer contains MCT oil.

23. A system for delivery of an active substance for release in the intestinal tract comprising a particle having a core containing an active substance, said core being encapsulated by a first layer of a coating material containing a prolamine and at least one of a plasticizer and a hydrophobic substance, said prolamine being in an amount of about 10% to 70% of the total weight of the core, said first coating layer being encapsulated by a second coating layer containing an enteric compound at about 5% to 70% of the sum of the weights of the core and first coating layer, and said second coating layer being encapsulated by an exterior coating layer of a coating material containing a prolamine and at least one of a plasticizer and a hydrophobic substance, said prolamine being in an amount of about 20% to 70% of the sum of the weights of the core and the first two coating layers.

24. A system according to claim 23, wherein prolamine is zein.

25. A system according to claim 24, wherein the zein has an ash content of not greater than about 2%
by weight.

26. A system according to claim 23, 24 or 25, wherein the enteric compound comprises at least one material selected from the group consisting of acids and esters of methacrylic copolymers.

27. A system according to claim 23, 24 or 25, wherein said first layer and said exterior layer both contain MCT oil.

28. A system according to claim 26, wherein said first layer and said exterior layer both contain MCT
oil.

29. A system according to claim 23, 24, 25 or 28, wherein the active substance is selected from the group consisting of analgesics, antibiotics, antidepressants, antivirals, antibodies, immuno-modulators, oncolytics, immunogens, hormones, vaccines, enzymes, nutrients and dietary fiber.

30. A system according to claim 23, 24, 25 or 28, wherein the active substance is a .beta.-lactam antibiotic.

31. A system according to claim 29, wherein said first layer and said exterior layer both contain MCT
oil.

32. A system according to claim 30, wherein said first layer and said exterior layer both contain MCT
oil.

33. A system according to claim 23, 24, 25, 28, 31 or 32, wherein said first layer contains about 10%
to 30% of the prolamine, by weight of the total weight of the core; said second coating contains about 20% to 40%, by weight, of the enteric compound, based on the sum of the total weights of the core and the first layer; and said exterior layer contains about 40% to 70%, by weight, of the sum of the weights of the core and the first and second layers.