US20070060542A9 - Method for preventing vascular disease - Google Patents

Abstract

A method for orally administering vitamin preparations is described which combine vitamin B₁₂ (B₁₂, cobalamin) and folic acid (folate), with and without pyridoxine (B₆), for preventing and treating elevated serum homocysteine (HC), cystathionine (CT), methylmalonic acid (MMA), or 2-methylcitric acid (2-MCA) levels. These metabolites have been shown to be indicative of B₁₂ and/or folic acid deficiencies. Further, it is likely that a B₆ deficiency may be present with a B₁₂ or folate deficiency. The method of the invention is also for use in lowering serum HC, CT, MMA, or 2-MCA in patients with or at risk for neuropsychiatric, vascular, renal or hematologic diseases. The method of the present invention eliminates the costly and time consuming steps of distinguishing between vitamin deficiencies once a deficiency is found by measurement of serum metabolite levels. The present invention is of particular benefit to the populations at risk for elevated serum metabolite levels, such as the people over the age of 65, and populations that have or are at risk for neuropsychiatric, vascular, renal and hematologic diseases.

Description

• This application is a continuation of Ser. No. 09/793,214, filed on Feb. 26, 2001, which is a continuation of Ser. No. 09/273,754 filed Mar. 22, 1999, now issued as U.S. Pat. No. 6,297,224, which is a continuation of application no. Ser. No. 09/012,955 filed Jan. 26, 1998 now issued as U.S. Pat. No. 5,795,873, which is a divisional of application no. Ser. No. 07/999,499, which was filed Dec. 29, 1992, now issued as U.S. Pat. No. 5,563,126.
FIELD OF THE INVENTION
• This invention relates to the field of nutrition. Specifically, the invention is comprised of new oral vitamin preparations combining vitamin B₁₂ (B₁₂, cobalamin) and folic acid (folate), and vitamin B₁₂, folate, and pyridoxine (B₆) for use in patients with elevated serum metabolite levels of homocysteine (HC), cystathionine (CT), methylmalonic acid (MMA), or 2-methylcitric acid (2-MCA). The elevation of these metabolites has been shown to be indicative of tissue deficiencies of B₁₂ and/or folate and/or B₆, and related to increased risk of neuropsychiatric, vascular, renal and hematologic diseases. One embodiment of the present invention uses a non-prescription formulation comprising between 0.3-10.0 mg B₁₂ and 0.1-0.4 mg folate, with the preferred embodiment using 2.0 mg B₁₂ and 0.4 mg folate. Another embodiment of the non-prescription formulation uses 0.3-10 mg B₁₂, 0.1-0.4 mg folate, and 5-75 mg B₆, with the preferred embodiment using 2.0 mg B₁₂, 0.4 mg folate, and 25 mg B₆. Another embodiment of the present invention uses a prescription strength formulation comprising between 0.3-10.0 mg B₁₂ and 0.4-1.0 mg folate, with the preferred embodiment using 2 mg B₁₂ and 1.0 mg folate. In a further embodiment of the present invention, a prescription strength formulation is used comprising 0.3-10 mg B₁₂, 0.4-1.0 mg folate, and 5-75 mg B₆, with the preferred embodiment using 2 mg B₁₂, 1.0 mg folate, and 25 mg B₆. The formulations of the present invention eliminate the costly and time-consuming steps of distinguishing between vitamin deficiencies once a deficiency is found by measurement of serum metabolite levels. The present invention is of particular benefit to the populations at risk for tissue deficiencies of B₁₂, folate, and B₆, such as people over the age of 65, and populations that have or are at risk for neuropsychiatric, vascular, renal and hematologic diseases.
BACKGROUND
• Vitamins B₁₂, folate, and B₆ are required cofactors in metabolic pathways involving methionine, homocysteine, cystathionine, and cysteine. B₁₂ in the form of 5′-deoxyadenosylcobalamin is an essential cofactor in the enzymatic conversion of methylmalonylCoA to succinylCoA. The remethylation of homocysteine (HC) to methionine catalyzed by methionine synthase requires folate (methyltetrahydrofolate) and B₁₂ in the form of methylcobalamin. HC is condensed with serine to form cystathionine (CT) in a reaction catalyzed by cystathionine □-synthase which requires B₆ (pyridoxal phosphate). CT is hydrolyzed in another B₆-dependent reaction to cysteine and □-ketobutyrate.
• It is important to diagnose and treat B₁₂, folate, and B₆ deficiencies because these deficiencies can lead to life-threatening hematologic abnormalities which are completely reversible by proper treatment. B₁₂ deficiency is a multisystem disorder with extremely varied clinical presentation which has been thought to occur in 0.4% of the population, e.g., about 1 million people in the United States. Symptoms of B₁₂ deficiency include significant anemia, displayed for example in decreased hematocrit (e.g., <25%) or hemoglobin (e.g., <8 g %), with macrocytic red blood cells (i.e., mean cell volume generally greater than 100 fl), or neurologic symptoms of peripheral neuropathy and/or ataxia. See, for example, Babior and Bunn (1983) in Harrison's Principles of Internal Medicine, (Petersdorf et al., eds.), McGraw-Hill Book Co., New York; Lee and Gardner (1984) in Textbook of Family Practice, 3rd Ed. (Rakel, ed.), Saunders & Co., Philadelphia). The hematological abnormalities seen are due to intracellular folate deficiency since folate is required for a number of essential enzymatic reactions involved in DNA and RNA synthesis and since the form of folate in serum (5-methyltetrahydrofolate) must be metabolized to tetrahydrofolate by the B₁₂-dependent enzyme methionine synthase before it can be utilized by the RNA- and DNA-related enzymes. While it has been well recognized that individuals with B₁₂ deficiency could display neurologic disorders in the absence of anemia, such situations were believed to be exceptional and rare. See, Beck (1985) in Cecil Textbook of Medicine, 17th Ed., (Wyngaarden and Smith, eds.), W. B. Saunders, Philadelphia, pp. 893-900; Babior and Bunn (1987) in Harrison's Principles of Internal Medicine, 11th Ed., (Braunwald et al., eds.) McGraw-Hill, New York, pp. 1498-1504; Walton (1985) in Brain's Diseases of the Nervous System, 9th Ed., Oxford University Press, Oxford, UK. The neurologic symptoms of B₁₂ deficiency were considered to be late manifestations of the disease most typically occurring after the onset of anemia or, if they occurred first, were soon to be followed by the onset of anemia. See, Woltmann (1919) Am. J. Med. Sci. 157:400-409 Victor and Lear (1956) Am. J. Med. 20:896-911.
• However, it has recently been shown that the textbook description of severe megaloblastic anemia and combined systems disease of the nervous system is the rarest presentation of B₁₂ deficiency at the present time (Stabler et al. (1990) Blood 76:871-881; Carmel (1988) Arch. Int. Med. 148:1712-1714 Allen (1991) in Cecil Textbook of Medicine, 19th Ed., (Wyngaarden and Smith, et al. eds.), W. B. Saunders, Philadelphia, pp. 846-854.). Therefore, contrary to previous teachings, patients that may benefit from B₁₂ therapy may have minimal to no hematologic changes while manifesting a wide variety of neurologic and psychiatric abnormalities (Lindenbaum et al. (1988) N. Engl. J. Med. 318:1720-1728; Greenfield and O'Flynn (1933) Lancet 2:62-63). This is particularly true for populations at risk for B₁₂ deficiency, such as the elderly population (Pennypacker et al. (1992) J. Am. Geriatric Soc. 40: (in press).
• The incidence of folate deficiency in the population is unknown, but has been thought to occur commonly in individuals with various degrees of alcoholism. The hematologic abnormalities seen with folate deficiency, such as macrocytic anemia, are indistinguishable from those seen with B₁₂ deficiency. Folate is required for a number of essential enzymatic reactions involved in DNA and RNA synthesis, and is particularly important in rapidly dividing cells like those in the bone marrow.
• B₆ is required for the first step in heme synthesis and serves a major role in transamination reactions of amino acid metabolism, in decarboxylations, and in the synthesis of the neuroactive amines histamine, tyramine, serotonin, and □-aminobutyric acid (GABA). Clinical manifestations include microcytic hypochromic anemia, characteristic skin changes of dermatitis and acrodynia, muscular weakness, and a variety of neuropsychiatric abnormalities including hyperirritability, epileptiform convulsions, depression and confusion (Newberne and Conner (1989) in Clinical Biochemistry of Domestic Animals, Academic Press, San Diego, pp. 796-834).
• Vitamin deficiencies are generally determined by measurement of serum levels. Normal serum B₁₂ levels are 200-900 pg/ml, with levels of less than 100 pg/ml being said to indicate clinically significant deficiency (Beck (1985) supra) However, serum B₁₂ levels are a relatively insensitive determinant of B₁₂ deficiency in that only 50% of patients with clinically confirmed B₁₂ deficiency have levels less than 100 pg/ml, 40% are 100-200 pg/ml, and at least 5-10% have values in the 200-300 pg/ml range. Diagnosis is further complicated by the fact that 2.5% of normal subjects (6,250,000 people in the U.S.) have low serum B₁₂ levels (Allen (1991) supra), with no evidence of B₁₂ deficiency and are unlikely to benefit from B₁₂ therapy (Schilling et al. (1983) Clin. Chem. 29:582; Stabler (1990) supra).
• Normal serum folate levels are 2.5-20 ng/ml, with levels less than 2.5 ng/ml indicating the possibility of clinically significant deficiency. Like B₁₂ serum levels, however, serum folate levels are a relatively insensitive measure in that only 50-75% of patients with folate deficiency have levels less than 2.5% ng/ml, with most of the remaining 25-50% being in the 2.5-5.0 ng/ml range (Allen (1991) in Cecil Textbook of Medicine, 19th Ed., su ra) The development of sensitive serum metabolite assays for HC, CT, MMA, and 2-MCA has allowed the relationship between metabolite levels and vitamin deficiencies to be investigated (Stabler et al. (1987) Anal. Biochem. 162:185-196; Stabler et al. (1986) J. Clin. Invest. 77:1606-1612; Stabler et al. (1988) J. Clin. Invest. 81:466-474). It has been found that elevated serum levels of HC and MMA are clinically useful tests of functional intracellular deficiencies of B₁₂ and folate, with elevated HC levels seen with both B₁₂ and folate deficiencies, and elevated MMA levels seen with a B₁₂ deficiency (Allen et al. (1990) Am. J. Hematol. 34:90-98 Lindenbaum et al. (1990) Am. J. Hematol. 34:99-107; Lindenbaum et al. (1988) N. Engl. J. Med. 318:1720-1728; Beck (1991) in Neuropsychiatric Consequences of Cobalamin Deficiency, Mosby Year Book 36:33-56 Moelby et al. (1990) 228:373-378; Ueland and Refsum (19890 J. Lab. Clin. Med. 114:473-501; Pennypacker et al. (1992) supra). Increased serum levels of CT are seen in both deficiencies and 2-MCA is elevated in B₁₂ deficiency (Allen et al. (1991) in Proceedings of the 1 st International Congress on Vitamins and Biofactors in Life Science, Kobe (Japan) ; Allen et al. (1993) Metabolism (in press)). HC and CT may be elevated in patients with intracellular deficiency of B₆, but this has not been as well documented (Park and Linkswiler (1970) J. Nutr. 100:110-116; Smolin and Benvange (1982) J. Nutr. 112:1264-1272).
• Elevated serum metabolite levels are observed in disease states other than classic vitamin deficiencies. For example, elevated HC levels have been observed in the presence of vascular disease. The homocysteine theory of atherosclerosis, formulated by McCully and Wilson (1975) Atherosclerosis 22:215-227, suggests that high levels of HC are responsible for the vascular lesions seen in homocystinuria, a genetic defect caused by a deficiency in the enzyme cystathionine □-synthase. The theory also implies that moderate elevations of HC might be associated with increased risk for vascular disease (Ueland et al. (1992) in Atherosclerotic Cardiovascular Disease, Hemostasis, and Endothelial Function (Francis, Jr., ed.), Marcel Dekker, Inc., New York, pp. 183-236). Moderate hyperhomocysteinemia has been shown to be frequently present in cases of stroke and to be independent of other stroke risk factors (Brattstrom et al. (1992) Eur. J. Clin. Invest. 22:214-221). Clinical and experimental evidence demonstrates that patients who are homozygotes for cystathionine □-synthase deficiency have a markedly increased incidence of vascular disease and thrombosis. A number of studies (see, Clarke et al. (1991) N. Engl. J. Med. 324:1149-1155) strongly suggest that heterozygotes for a deficiency of cystathionine β-synthase also have an increased incidence of vascular disease and thrombosis and that such heterozygotes may constitute as many as one-third of all patients who develop strokes, heart attacks, or peripheral vascular disease under age 50. It is also likely that such heterozygotes are also at increased risk for vascular disease and thrombosis after age 50. Since the incidence of heterozygosity for cystathionine β-synthase deficiency is estimated to be 1 in 60-70, this means that there are approximately 4 million heterozygotes in the U.S. It is also possible that patients with vascular disease due to other causes, such as hypercholesterolemia, would also benefit from a decrease in their serum HC levels even if their existing levels are only slightly elevated or actually within the normal range.
• Renal disease is another condition that gives rise to elevated levels of serum metabolites. Approximately 75% of patients with renal disease have elevated serum concentrations of HC, CT, MMA, and 2-MCA. Since patients with renal disease have a significant incidence and marked acceleration of vascular disease, it might be beneficial to lower their serum metabolite levels, especially that of HC.
• An increasing prevalence of low serum B₁₂ concentrations with advancing age has been found by many but not all investigators (Bailey et al. (1980) J. Am. Geriatr. Soc. 28:276-278 Eisborg et al. (1976) Acta Med. Scand. 200:309-314; Niisson-Ehle et al. (1989) Dig. Dis. Sci. 34:716-723; Norman (1985) 33:374; Hitzhusen et al. (1986) Am. J. Clin. Pathol. 85:3236), folate (Magnus et al. (1982) Scan. J. Haematol. 28:360-366; Blundell et al. (1985) J. Clin. Pathol. 38:1179-1184 Elwood et al. (1971) Br. J. Haematol. 21:557-563; Garryet al. (1984) J. Am. Geriatr. Soc. 32:71926; Hanger et al. (1991) J. Am. Geriatr. Soc. 39:1155-1159), and B₆ (Ranke et al. (1960) J. Gerontol. 15:41-44; Rose et al. (1976) Am. J. Clin. Nutr. 29:847-853; Baker et al. (1979) J. Am. Geriatr. Soc. 27:444-450). Moreover, prevalence estimates for these vitamin deficiencies vary widely depending on the population groups studied. It has been unclear whether this increased prevalence is a normal age related phenomena or a true reflection of tissue vitamin deficiency and whether the low serum vitamin concentrations are a reliable indicator of functional intracellular deficiency.
• It is difficult, expensive and time-consuming to distinguish between deficiencies of vitamins B₁₂, folate, and B₆. The hematologic abnormalities seen with B₁₂ deficiency are indistinguishable from those seen with folate deficiency. Similarly to a B₁₂ deficiency, B₆ deficiencies also result in hematologic as well as neuropsychiatric abnormalities. The traditional methods of determining deficiencies by measurement of serum vitamin levels are often insensitive. As a result, in order to determine if and which vitamin deficiency is present, a patient will be treated with one vitamin at a time and the response to that vitamin determined by normalization of serum vitamin levels and the correction of hematologic abnormalities. These steps are then repeated with each vitamin. This method of treatment is both expensive and time-consuming. In the presence of multiple deficiencies, the diagnosis of vitamin deficiencies is further confused and give rise to the dangerous possibility that only one deficiency will be treated. For example, the hematologic abnormalities seen with a B₁₂ deficiency will respond to treatment with folate alone. However, the neuropsychiatric abnormalities caused by the B₁₂ deficiency will not be corrected and may indeed by worsened.
• It has now been discovered for the first time that the prevalence of intracellular deficiencies of vitamins B₁₂, folate, and B₆, alone or in combination, is substantially higher than that previously estimated by measurement of serum vitamin concentrations. The present disclosure establishes that tissue deficiencies of one or more of the vitamins B₁₂, folate and B₆, as demonstrated by the elevated metabolite concentrations, occurs commonly in the elderly population even when serum vitamin levels are normal. Based on this new discovery, the present invention addresses the problem of distinguishing between vitamin deficiencies when low, low-normal, or normal serum vitamin concentrations are found by providing formulations for the treatment of high serum metabolites and at-risk populations for combinations of one or more tissue deficiencies of vitamins B₁₂, folate, and B₆.
• Hathcock and Troendle (1991) JAMA 265:96-97, have suggested the treatment of pernicious anemia with an oral pill containing 300 to 1000 ug or more per day of B₁₂. However, contrary to the present invention, Hathcock and Troendle teach away from combining B₁₂ therapy with folate, since “if the oral cobalamin therapy should fail to maintain adequate levels, folate might provide protection against development of anemia while permitting nerve damage from cobalamin deficiency.”
• U.S. Pat. No. 4,945,083, issued Jul. 31, 1990 to Jansen, entitled: Safe Oral Folic-Acid-Containing Vitamin Preparation, describes a oral vitamin preparation comprising 0.1-1.0 mg B₁₂ and 0.1-1.0 mg folate for the treatment or prevention of megaloblastic anemia. This formulation presents a problem in the case of a B₁₂ deficient patient, in that the 0.5 mg folate may correct the hematologic abnormalities present, but the 0.5 mg B₁₂ dose may be insufficient to correct a B₁₂ deficiency due to inadequate intrinsic factor. By contrast, the formulation of the present invention teaches the use of the combination of B₁₂ and folate, and of B₁₂, folate and B₆, sufficient to treat either single or multiple deficiencies of B₁₂, folate, and B₆. The present invention does not rely on the determination of vitamin deficiencies by the measurement of serum vitamin levels, but uses the more sensitive measurement of elevated serum metabolites of HC, CT, MMA, and 2-MCA, shown to be related to the presence of B ₁₂ and/or folate and/or to B6 deficiencies or to the presence of the increased risk of neuropsychiatric, vascular, renal, and hematologic diseases.
• It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention as claimed.
SUMMARY OF THE INVENTION
• This invention includes a method for orally administering two new vitamin preparations containing vitamin B₁₂ and folate, and vitamin B₁₂, folate and B₆, for the treatment of patients with elevated serum metabolites, such as homocysteine, cystathionine, methylmalonic acid, and 2-methylcitric acid, as well as populations at risk for tissue deficiencies in one or more of the vitamins B₁₂, folate, and B₆ or for neuropsychiatric, vascular, renal, or hematologic diseases.
• One embodiment of the present invention uses an over-the-counter formulation comprised of between 0.3-10 mg CN-cobalamin (B₁₂) and 0.1-0.4 mg folate. Another embodiment of the non-prescription formulation uses 0.3-10 mg B₁₂, 0.1-0.4 mg folate, and 5-75 mg B₆. Preferred embodiments of the over-the-counter formulation are comprised of about 2.0 mg B₁₂ and 0.4 mg folate, and 2.0 mgB₁₂, 0.4 mg folate, and 25 mg B₆, respectively.
• Another embodiment of the present invention uses a prescription formulation comprised of between 0.3-10 mg CN-cobalamin (B₁₂) and 0.4-10.0 mg folate. Another embodiment of the prescription formulation of the present invention uses 0.3-10 mg B₁₂, 0.4-10.0 mg folate, and 5-75 mg B₆. Preferred embodiments of the prescription formulation use about 2.0 mg B₁₂ and 1.0 mg folate, and 2.0 mg B₁₂, 1.0 mg folate, and 25 mg B₆, respectively.
BRIEF DESCRIPTION OF THE DRAWINGS
• FIG. 1 shows the distribution of serum B₁₂ levels for a population of elderly outpatients (ages 65-99, n=152) and a normal population (ages 17-65, n=100).
• FIG. 2 shows serum MMA levels for a population of elderly outpatients with serum B₁₂ values <300 pg/ml (ages 65-99, n=38/152) and a normal population with serum B₁₂ values <300 pg/ml (ages 17-65, n=10/100).
• FIG. 3 shows serum HC levels for a population of elderly outpatients with serum B₁₂ values <300 pg/ml (ages 65-99, n=38/152) and a normal population with serum B,₂ values <300 pg/ml (ages 17-65, n=10/100).
• FIG. 4 shows serum MMA levels before and after treatment with parenteral cobalamin for a population of elderly outpatients with elevated MMA values and serum B₁₂ values <300 pg/ml (ages 65-99, n=15/38).
• FIG. 5 shows serum HC levels before and after treatment with parenteral cobalamin for a population of elderly outpatients with elevated HC values and serum B₁₂ values of <300 pg/ml (ages 65-99, n=10/38).
• FIG. 6 shows the distribution of serum B₁₂ levels for a population of elderly nursing home patients (ages 55-107, n=212) and a normal population (ages 17-65, n=100).
• FIG. 7 shows serum MMA levels for a population of elderly nursing home patients with serum B₁₂ values <300 pg/ml (ages 55-107, n=29/212) and a normal population with serum B₁₂ values (ages 17-65, n=10/100).
• FIG. 8 shows serum HC levels for a population of elderly nursing home patients with serum B₁₂ values <300 pg/ml (ages 55-107, n=29/212) and a normal population with serum B12 values <300 pg/ml (ages 17-65, n=10/100).
• FIG. 9 shows serum MMA levels before and after treatment with parenteral cobalamin for a population of elderly nursing home patients with serum B₁₂ values <300 pg/ml (ages 55-107, n=14/29).
• FIG. 10 shows serum HC levels before and after treatment with parenteral cobalamin for a population of elderly nursing home patients with serum B₁₂ values <300 pg/ml (ages 55-107, n=14/29).
• FIG. 11 shows the distribution of serum B₁₂ levels for a population of elderly patients (ages 65-99, n=548) and a normal population (ages 22-63, n=1 17) (Framingham study).
DETAILED DESCRIPTION OF THE INVENTION
• Reference will now be made in detail to the presently preferred embodiments of the invention, which, together with the following examples, serve to explain the principles of the invention.
• This invention uses new oral vitamin formulations combining vitamin B₁₂ (B₁₂, cobalamin) and folic acid (folate), and vitamin B₁₂, folate and pyridoxine (B₆). The formulations of the present invention are for use in the treatment of elevated serum levels of one or more of the metabolites homocysteine (HC), cystathionine (CT), methylmalonic acid (MMA), or 2-methylcitric acid (2-MCA). The use of the formulations of the present invention further include as a method of lowering serum metabolite levels of one or more of HC, CT, MMA, or 2-MCA, where these metabolite levels are not elevated but the patients are at risk for or have neuropsychiatric, vascular, renal, or hematologic diseases.
• One embodiment of the present invention uses a non-prescription formulation comprised of between about 0.3-10 mg CN-cobalamin (B₁₂) and 0.1-0.4 mg folate. Another embodiment of the present invention uses a non-prescription formulation comprised of between about 0.3-10 mg B₁₂, 0.1-0.4 mg folate, and 5-75 mg B₆. Preferred embodiments of the non-prescription formulation are comprised of about 2.0 mg B₁₂ and 0.4 mg folate, and 2.0 mg B₁₂, 0.4 mg folate, and 25 mg B₆, respectively.
• Another embodiment of the present invention is comprised of a prescription formulation comprised of between about 0.3-10 mg B₁₂ and 0.4-10.0 mg folate, with the preferred embodiment comprised of about 2.0 mg B₁₂ and 1.0 mg folate. Another embodiment of the prescription strength formulation is comprised of about 0.3-10 mg B₁₂, 0.4-10.0 mg folate, and 5-75 mg B₆, with a preferred embodiment comprised of about 2.0 mg B₁₂, 1.0 mg folate, and 25 mg B₆.
• The formulations of the present invention are for the treatment and prevention of elevated metabolite levels in at risk populations, such as the elderly, and people that have or are at risk for neuropsychiatric, vascular, renal and hematologic diseases. The present invention eliminates the costly and time consuming need to differentiate between B₁₂, folate, and B₆ deficiencies.
• The administration of a daily dose of the vitamin formulations of the present invention provides better long-term normalization of serum HC and other metabolites than prior art formulations, and eliminates the difficulty in differentiating between deficiencies of two or three of the vitamins, the difficulty in diagnosing multiple deficiencies of two or three of the vitamins, and the expense of doing so. Further, the administration of an oral preparation of B₁₂ and folate, with or without B₆, is preferred over intramuscular injections for patient convenience and ease of administration.
• For example, the inclusion of B₁₂ will be useful as a safeguard for patients misdiagnosed as folate deficient, even though they are actually B₁₂ deficient, since treatment with folate alone in such patients is extremely dangerous. The danger arises from the fact that treating a B₁₂ deficient patient with folate alone may reverse or prevent the hematologic abnormalities seen in B₁₂ deficiency, but will not correct the neuropsychiatric abnormalities of a B₁₂ deficiency and may actually precipitate them. Even in the absence of intrinsic factor, approximately 1% of a 2.0 mg oral dose of B₁₂ is absorbed by diffusion. Thus, approximately 20 ug of B₁₂ would be absorbed from the formulations of the present invention which would be more than adequate even in patients with pernicious anemia who have lost their intrinsic factor-facilitated absorption mechanism for B₁₂. The inclusion of folate will be of benefit since B₁₂ deficiency causes a secondary intracellular deficiency of folate. The inclusion of folate and B₆ will also be of benefit in patients with mixed vitamin deficiencies.
• The formulations of the present invention may be administered as a non-injectable implant or orally. Non-injectable use may be as a patch. Formulations for oral administration are preferably encapsulated. Preferably, the capsule is designed so that the formulation is released gastrically where bioavailability is maximized. Additional excipients may be included to facilitate absorption of the vitamin formulations. Diluents, flavorings, low melting point waxes, vegetable oils, lubricants, suspending agents, tablet disintegrating agents, and binders may also be employed.
• Example 1 describes the methods used to measure serum vitamin and metabolite levels. Example 2 describes a new study conducted with 412 subjects over the age of 65 with a variety of medical conditions correlating the incidence of low serum vitamin levels with elevated serum metabolite levels. A study determining the incidence of undetected B12 deficiency and response of serum MMA and HC to B₁₂ treatment in a geriatric outpatient population is described in Example 3. Example 4 describes a similar study conducted with a geriatric nursing home population, and Example 5 describes a similar study conducted with another geriatric population.
EXAMPLE 1
• Methods for Measurement of Serum Vitamin and Metabolite Levels.
• Serum vitamin assays. Serum vitamins B12 and folate were measured by a quantitative radioassay method using purified intrinsic factor and purified folate binding protein. Vitamin B6 was measured by a radioenzymatic assay method wherein serum is incubated with apoenzyme tyrosine-decarboxylase, C₁₄ labelled tyrosine is added to start the enzymatic reaction which is stopped with HCl. Subsequently the free C₁₄-labelled CO₂ is adsorbed by a KOH impregnated filtering paper. The measured C₁₄ activity is directly proportional to the B₆ (pyridoxal phosphate) concentration (Laboratory Bioscientia, Germany).
• Serum metabolite assays. Serum metabolite assays for homocysteine and methylmalonic acid were conducted by the capillary gas chromatography and mass spectrometry methods of Marcell et al. (1985) Anal. Biochem. 150:58; Stabler et al. (1987) supra, and Allen et al. (1990) Am. J. Hematol. 34:90-98. Serum cystathionine levels were assayed by the method of Stabler et al. (1992) Blood (submitted). Serum 2-methylcitric acid was assayed by the method of Allen et al. (1993) Metabolism supra.
• Statistical methods. Statistical analysis was done with the SAS statistical package (version 6.06). Nonparametric data for two or more groups were tested with the two sample Wilcoxon rank sum test (with Bonferroni's correction for the significance level ax) and the Kruskall Wallis test. From the results of the healthy young subjects reference intervals were calculated. Since the frequency distribution of the values of each parameter were markedly abnormal they were transformed to normal distributions using log transformation. The sample prevalence p with 95% confidence intervals of low serum vitamins B₁₂, folate, and B₆ concentrations was calculated as (p±2p (1−p)/n×100 wherein n is the total sample size, p is the number of low serum vitamin concentrations/n; low serum concentrations are defined as <mean −2 S.D.
EXAMPLE 2
• Incidence of Elevated MMA, 2-MCA, HC, and CT Levels in the Geriatric Population.
• The serum concentrations of B₁₂, folate, and B₆ were measured in 412 subjects over the age of 65 (subgroups A-D), and in 99 healthy control subjects aged 20-55 years (subgroup E). The geriatric subgroups were defined as follows: A, 110 patients with atherosclerosis; B, 98 patients with neuropsychiatric disorders; C, 102 patients with atherosclerosis and multiple diseases including rheumatoid arthritis and diabetes; D, 102 subjects who were healthy.
• Venous blood was obtained from all subjects in the morning after an overnight fast. The blood was spun within one hour after collection and the serum was transported in dry ice to the central laboratory. Serum vitamins B₁₂ and folate were measured as described in Example 1 with a vitamin B₁₂/folate dual RIA kit (CT301/CT302 Amersham Buchier, UK). Vitamin B₆ and serum metabolites were measured as described in Example 1.
• Since renal function can influence serum metabolite concentrations (Ueland and Refsum (1989) supra Moelby et al. (1992) Scand. J. Clin. Lab. Invest. 52:351-354), serum creatinine concentrations were measured in all subjects by the Jaffe photometric method (Laboratory Bioscientia, Germany). Normal range was 62-124 μmol/L. Creatinine clearance was calculated using the formulation of Cockroft and Gault (1976) Nephron 16:31-41.
• Normal ranges for serum vitamin and metabolite levels were determined by the mean ±2 standard deviations after log normalization using the values from subgroup E. Results are shown in Table 1:

  TABLE 1
  INCIDENCE OF LOW SERUM
  VITAMN AND HIGH METABOLITE LEVELS
  IN GERIATRIC POPULATIONS A-D AND
  A YOUNGER HEALTHY POPULATION E.

  		Folic
  Group	B12	Acid	B6	MMA	2-MCA	HC	CT
  A	6%	12%	48%	36%	44%	55%	64%
  B	6%	19%	53%	47%	39%	59%	6%
  C	3%	10%	50%	32%	45%	39%	73%
  D	6%	6%	17%	26%	23%	38%	41%
  E	2%	1%	1%	3%	6%	2%	4%

• There was a rough correlation with low vitamin levels and elevated metabolites, but many of the patients with elevated metabolites had low normal or normal vitamin levels. Correlations between clinical abnormalities within groups A, B, and C were not present. Patients were treated with weekly injections of a multi-vitamin preparation containing 1.0 mg B₁₂, 1.1 mg folate, and 5 mg B₆, resulting in a marked lowering or normalization of elevated metabolite levels in virtually every elderly patient.
• These data support the conclusions that there is an increased incidence of low levels of serum B₁₂, folate, and B₆ in the geriatric population, and that serum MMA, 2-MCA, HC and CT are elevated in an even higher percentage of geriatric patients. The presence of elevated levels of one or more of the metabolites HC, CT, MMA, or 2-MCA indicate a tissue or intracellular deficiency of one or more of the vitamins B₁₂, folate and B₆. It not possible to tell without expensive, time-consuming, and extensive testing which one vitamin or pair of vitamins, or whether all three vitamins are deficient. These observations, together with the fact that elevated metabolite levels are corrected by parenteral therapy with a combination of vitamins B₁₂, folate, and B₆, indicate that a tissue deficiency of one or more of these vitamins occurs commonly in the geriatric population and that measurement of serum vitamin levels alone is an inadequate method for identifying such deficiencies.
EXAMPLE 3
• Determination of Serum B₁₂ Folate, MMA, HC, CT and 2-MCA Levels in a Geriatric Outpatient Population.
• A study was conducted with 152 elderly outpatient subjects to measure the prevalence of B₁₂ deficiency in geriatric outpatients as determined by both low serum B₁₂ levels and elevations of MMA and HC, and to determine the response to B₁₂ treatment. Blood samples were obtained on 152 consecutive geriatric outpatients, ages 65-99. Control values were determined from 100 subjects, ages 17-65. Serum B₁₂ folate, MMA, HC, CT, and 2-MCA levels were obtained for each patient, shown in Table 2. The significance of the results marked as “**” in Table 2 are as follows: B₁₂ levels of <200 pg/ml; folate <3.8 ng/ml; homocysteine >16.2 uM; MMA >271 nM; CT >342 nM; and 2-MCA >228 nM. Serum MMA, HC, CT, and 2-MCA levels were measured as described in Example 1. Serum B₁₂ and folate were measured as described in Example 1 using a Coming humophase kit (CIBA-Corning, Medfield, Mass.) with the normal range defined as 200-800 pg/ml for B₁₂ and 3.8 ng/ml for folate. After evaluation, patients received weekly parenteral cyanocobalamin injections (1,000 ug IM) for 8 weeks, followed by monthly injections. Repeat laboratory and clinical assessments were administered at 8 weeks and at 6 months.
• Results show that 25% of the subjects had a serum B₁₂ level <300 pg/ml and 8.5% had a low level of <200 pg/ml. FIG. 1 shows the shift seen in elderly subject towards lower serum B₁₂ levels. More than half of the subjects with low or low-normal serum B₁₂ levels had elevations of MMA (FIG. 2) and/or HC (FIG. 3) greater than 3 S.D. above the means in normals and representing 14.5% of the total screened population.
• Patients with low and low/normal serum B₁₂ levels were treated with weekly injections of 1.0 mg B₁₂. Parenteral B₁₂ administration caused elevated metabolite levels to fall to or towards normal (FIGS. 4 and 5) in every subject treated with B₁₂. It appears that the true prevalence of previously unrecognized B₁₂ deficiency in this elderly population was at least 14.5%.
• It can be seen from the data presented in Table 2 that serum B₁₂ levels are insensitive for screening B₁₂ deficiencies since similar numbers of patients with low normal serum B₁₂ levels of 201-300 pg/ml compared with patients with low B₁₂ levels (<200 pg/ml) had markedly elevated metabolites which fell with B₁₂ treatment. Further, this study shows that elderly patients have a high incidence (at least 14.5%) of unrecognized B₁₂ deficiency, detectable by measurement of serum HC and MMA levels in patients with serum B₁₂ levels <300 pg/ml.
• A further finding in this study emphasizes the need to treat elevated metabolite levels with a combination of vitamin B₁₂ and folate with or without B₆. Some of the patients exhibiting elevated metabolite levels did not fully respond to B₁₂ treatment. This may indicate a concomitant deficiency of folate and/or B₆. The lack of response to B₁₂ treatment could result from a deficiency of one, a pair, or all three vitamins. However, it would be expensive and time-consuming to attempt to distinguish between the vitamin deficiencies.
• Another, and perhaps the most important, finding in this study is the large number of patients with serum B₁₂>300 pg/ml who have elevated values for one or more metabolites as indicated by a “**” next to the individual values. As can readily be seen in Table 2, there are many examples of elevated value for MMA and/or 2-MCA at all levels of serum B₁₂ including the mid-normal (300-600 pg/ml), the high-normal (600-800 pg/ml), and even the elevated (>800 pg/ml) ranges. The same is true for elevations of HC and CT. In some patients the serum folate is low, indicating that folate deficiency may be present, but in many cases both B₁₂ and folate levels are normal. B₆ levels were not performed in this study, but B₆ deficiency would not be expected to cause elevations of MMA or 2-MCA. Thus in many patients it is not clear which vitamin, or pair of vitamins, or whether all three vitamins is or are deficient. One could pick a single vitamin, often at random, with which to treat a patient for several weeks or months, and then repeat measurement of metabolite levels to determine if a partial or full correction had occurred. If there was no response, one could try another vitamin, or if there was a partial response one could add a second vitamin, and then repeat metabolite measurement after several weeks or months. If there was still no response, one could try the third vitamin, or if there was a partial response, one could try a different pair of vitamins. Eventually one could determine whether an individual vitamin, a particular pair of vitamins, or all three vitamins were required to normalize or maximally reduce the metabolite levels, but it would often require months or even a year to make this determination. Such a determination would be expensive. In addition, a patient who was optimally treated with a single vitamin or pair of vitamins might subsequently develop a deficiency of one or even two of the other vitamins as evidenced by a re-elevation or increase in the levels of one or more metabolites. Therapeutic testing could be reinitiated and continued as described above, although this would also be time-consuming and expensive.
• It requires less time and expense to treat patients with elevated metabolite levels with a combination of vitamin B₁₂ and folate, or a combination of vitamin B₁₂, folate and vitamin B₆. The utility of the approach of the present invention is appreciated only after it is taught, for the first time in the present disclosure, that a deficiency of one or more of the three vitamins occurs commonly in the elderly population as evidenced by elevation of one or more metabolites, i.e., MMA, 2-MCA, HC and CT.
EXAMPLE 4
• Determination of Serum B₁₂, Folate, MMA, and HC Levels in a Geriatric Nursing Home Population.
• A study was conducted with 212 elderly nursing home patients to determine serum B₁₂, folate, MMA, and HC levels (Table 3). The significance of the results shown in Table 3 marked with “**” are as described for Table 2 (Example 3). The control group consisted of 100 subjects between the ages of 17-65 years. As in the study described in Example 3, the elderly population exhibited a shift to lower serum B₁₂ levels (FIG. 6), elevated serum MMA (FIG. 7) and HC (FIG. 8) levels. Parenteral administration of B ₁₂ 1 mg per week for 8 weeks to those with serum B₁₂<300 pg/ml caused elevated MMA (FIG. 9) and HC (FIG. 10) levels to fall to or towards normal.
• As in the study reported in Example 3, a further finding in this study emphasizes the need to treat elevated metabolite levels with a combination of vitamins B₁₂ and folate, with or without B₆. Some of the patients exhibiting elevated metabolite levels did not fully respond to B₁₂ treatment. This may indicate a concomitant deficiency of folate and/or B₆. The lack of response to B₁₂ treatment could result from a deficiency of one, a pair, or all three vitamins. However, it would be expensive and time-consuming to attempt to distinguish between the vitamin deficiencies.
• Again, an important finding in this study is the large number of patients with serum B₁₂>300 pg/ml who have elevated values for one or more metabolites as indicated by a “**” next to the individual values. As is seen in Table 3, there are many examples of elevated values for MMA at all levels of serum B₁₂ including the mid-normal (300-600 pg/ml), the high-normal (600-800 pg/ml), and even the elevated (>800 pg/ml) ranges. The same is true for elevations of HC. In some patients the serum folate is low, indicating that folate deficiency may be present, but in many cases both B₁₂ and folate levels are normal. B₆ levels were not performed in this study, but B₆ deficiency would not be expected to cause elevations of MMA. Thus, again it is not clear which vitamin, or pair of vitamins, or whether all three vitamins is or are deficient. One could pick a single vitamin with which to treat a patient for several weeks or months, and then repeat measurement of metabolite levels to determine if a partial or full correction had occurred. If there was no response, one could try another vitamin, or if there was a partial response one could add a second vitamin, and then repeat metabolite measurement after several weeks or months. If there was still no response, one could try the third vitamin, or if there was a partial response, one could try a different pair of vitamins. Eventually one could determine whether an individual vitamin, a particular pair of vitamins, or all three vitamins were required to normalize or maximally reduce the metabolite levels, but it would often require months or even a year to make this determination. Such a determination would be expensive. In addition, a patient who was optimally treated with a single vitamin or pair of vitamins might subsequently develop a deficiency of one or even two of the other vitamins as evidenced by a re-elevation or increase in the levels of one or more metabolites. Therapeutic testing could be reinitiated and continued as described above, although this would also be time-consuming and expensive.
• It requires less time and expense to treat patients with elevated metabolite levels with a combination of vitamin B₁₂ and folate, or a combination of vitamin B_12, folate and vitamin B₆. The utility of the approach of the present invention is appreciated only after it is taught, for the first time in the present disclosure, that a deficiency of one or more of the three vitamins occurs commonly in the elderly population as evidenced by elevation of one or more metabolites, i.e., MMA, 2-MCA, HC and CT.
EXAMPLE 5
• Determination of Serum B₁₂ Folate, I4MA, and HC Levels in a Geriatric Population.
• A study was conducted with 548 elderly subjects from the Framingham study between the ages of 65-99 to determine serum B₁₂, folate, MMA, and HC levels (Table 4). The significance of the results shown in Table 4 (marked with “**”) are as described for Table 2 (Example 2).
• As in the study described in Examples 3 and 4, the elderly population exhibited a shift to lower serum B₁₂ levels (FIG. 11), and elevated serum MMA and HC levels. The elderly population also exhibited a high incidence (9.5%) of low serum folate levels (Table 4). As in the studies reported in Examples 2, 3 and 4, the incidence of tissue or intracellular vitamin deficiencies based on elevated metabolite levels was higher than that predicted from measurement of serum vitamin levels.
• As in Examples 3 and 4 above, these results confirm the importance of the finding that there are a large number of patients with serum B₁₂>300 pg/ml who have elevated values for one or more metabolites as indicated by a “**” next to the individual values. As is seen in Table 4, there are many examples of elevated MMA values at all levels of serum B₁₂ including the mid-normal (300-600 pg/ml), the high-normal (600-800 pg/ml), and even the elevated (>800 pg/ml) ranges. The same is true for elevations of HC. In some patients the serum folate is low, indicating that folate deficiency may be present, but in many cases both B₁₂ and folate levels are normal. B₆ levels were not performed in this study, but B₆ deficiency would not be expected to cause elevations of MMA. Thus, again it is not clear which vitamin, or pair of vitamins, or whether all three vitamins is or are deficient. One could pick a single vitamin with which to treat a patient for several weeks or months, and then repeat measurement of metabolite levels to determine if a partial or full correction had occurred. If there was no response, one could try another vitamin, or if there was a partial response one could add a second vitamin, and then repeat metabolite measurement after several weeks or months. If there was still no response, one could try the third vitamin, or if there was a partial response, one could try a different pair of vitamins. Eventually one could determine whether an individual vitamin, a particular pair of vitamins, or all three vitamins were required to normalize or maximally reduce the metabolite levels, but it would often require months or even a year to make this determination. Such a determination would be expensive. In addition, a patient who was optimally treated with a single vitamin or pair of vitamins might subsequently develop a deficiency of one or even two of the other vitamins as evidenced by a re-elevation or increase in the levels of one or more metabolites. Therapeutic testing could be reinitiated and continued as described above, although this would also be time-consuming and expensive.
• It requires less time and expense to treat patients with elevated metabolite levels with a combination of vitamin B₁₂ and folate, or a combination of vitamin B_12, folate and vitamin B₆. The utility of the approach of the present invention is appreciated only after it is taught, for the first time in the present disclosure, that a deficiency of one or more of the three vitamins occurs commonly in the elderly population as evidenced by elevation of one or more metabolites, i.e., MMA, 2-MCA, HC and CT.

  TABLE 2
  SERUM METABOLITE & VITAMIN LEVELS IN A GERIATRIC
  OUTPATIENT POPULATION

  						Total
  Patient	B12	Folate	Homocysteine	MMA	CT	MC

  116	66**	9.8	41.8**	1508**	507**	759**
  118	79**	9.3	29.6**	2200**	343**	428**
  016	155**	7.6	15.3	1316**	208	196
  067	163**	6.6	9.9	93	164	69
  091	178**	12.0	29.2**	3108**	438**	318**
  042	181**	11.3	13.0	452**	300	262**
  030	185**	6.6	26.0**	282**	310	223
  037	187**	9.4	12.3	160	218	334**
  100	187**	9.5	13.6	208	453**	141
  036	188*	9.9	16.3**	298**	385**	322**
  109	189**	7.6	12.3	127	188	161
  007	191**	11.7	67.1**	6349**	619**	1005**
  018	193**	5.8	16.7**	412**	272	235**
  050	210	4.0	25.3**	464**	727**	121
  108	214	6.0	31.1**	264	523**	315**
  041	216	7.2	19.1**	418**	360**	288*
  126	224	6.5	8.8	103	361**	121
  005	231	12.5	17.1**	269	825**	276**
  024	235	13.0	18.5**	2946**	232	289**
  111	237	6.3	14.6	135	380**	203
  023	239	4.1	21.9**	385**	775**	279**
  010	256	12.9	11.5	652**	119	144
  055	258	6.8	7.5	189	342	185
  102	259	10.9	23.9**	1894**	423**	400**
  026	260	18.5	20.4**	1949**	295	248**
  107	262	13.1	10.1	231	628**	153
  038	269	7.6	15.7	222	152	152
  140	277	4.0	29.1**	744**	602**	254**
  074	278	5.2	24.1**	699**	296	187
  002	278	14.6	14.8	554**	259	277**
  019	282	8.5	12.4	329**	262	161
  035	287	5.8	9.8	230	390**	218
  049	290	3.9	33.0**	140	275	138
  078	290	10.9	12.5	197	240	209
  045	291	8.7	9.5	162	613**	132
  092	294	14.9	19.3**	500**	246	167
  137	297	6.8	10.1	631**	340	184
  072	298	6.7	19.7**	375**	302	246**
  149	310	8.3	16.1	314**	199	149
  047	312	4.9	15.9	277**	271	173
  060	312	9.4	8.0	100	228	203
  046	314	6.5	16.2	142	336	125
  093	318	6.4	16.5**	304**	361**	130
  014	321	14.5	10.7	275**	233	170
  088	327	7.1	17.8**	263	507**	258**
  032	340	6.6	8.6	150	133	133
  147	347	7.6	18.2**	305**	219	265**
  001	351	4.7	20.8**	199	402**	223
  090	353	4.9	20.7**	144	419**	178
  008	358	5.4	11.6	372**	529**	177
  104	360	12.7	12.1	260	89	77
  110	370	3.0**	17.1**	456**	297	150
  103	371	18.7	14.5	257	219	180
  056	373	6.5	12.4	236	415**	189
  048	374	3.6**	9.7	167	237	230**
  131	377	10.9	13.6	256	220	85
  122	378	76	21.9**	906**	227	196
  004	385	8.6	10.3	109	288	92
  120	390	9.8	22.9**	499**	529**	260**
  138	405	6.9	14.7	334**	238	188
  141	407	8.1	14.3	168	259	263**
  101	408	5.9	9.2	160	134	40
  145	410	3.7**	25.4**	567**	550	349**
  027	415	11.1	10.6	169	278	164
  028	418	5.6	34.6**	608**	589**	351**
  011	420	10.6	18.8**	683**	1014**	282**
  081	421	6.6	16.5**	861**	641**	531**
  033	423	4.2	16.3**	156	194	170
  057	425	18.3	13.5	209	381**	321**
  021	427	18.9	12.1	223	524**	168
  135	430	8.8	13.5	284**	412**	180
  097	435	15.4	10.9	353**	465**	119
  052	438	6.8	15.2	281**	372**	238**
  132	448	12.6	16.8**	1931**	394**	250**
  086	451	12.1	6.6	139	208	107
  148	458	13.9	11.4	187	322	238**
  012	466	15.3	8.3	560**	250	144
  083	466	12.0	13.7	366**	214	193
  133	470	13.8	10.8	290**	275	55
  017	475	4.0	39.6**	196	467**	220
  053	476	13.4	12.3	226	206	125
  009	482	6.5	25.3**	240	470**	214
  066	498	9.6	12.9	374**	233	92
  031	507	11.0	14.8	173	278	220
  099	507	10.4	9.6	124	233	63
  128	507	4.6	9.4	294**	324	176
  013	514	11.3	15.9	163
  151	522	7.8	14.3	370**	324	215
  077	523	6.8	17.7**	184	210	214
  079	523	15.6	13.0	316**	223	251**
  054	524	4.9	10.0	148	230	123
  020	524	9.9	14.2	235	366**	190
  069	528	7.0	9.7	257	281	83
  085	536	4.0	22.5**	97	191	114
  084	551	14.2	12.5	166	179	131
  082	559	12.3	14.6	208	371**	182
  117	560	3.4**	18.8**	102	176	88
  061	561	12.7	9.8	170	404**	152
  006	567	4.6	16.8**	138	688**	165
  129	567	4.9	16.2	363**	495**	331**
  003	570	11.4	12.9	189	330	230**
  115	576	6.3	17.8**	128	231	95
  089	578	10.3	12.0	147	258	236**
  143	581	2.6**	42.7**	165	555**	208
  114	583	5.1	16.6**	599**	660**	177
  080	593	9.5	18.0**	208	289	142
  015	598	7.0	12.4	167	381**	95
  039	598	9.6	18.1**	691**	719**	354**
  070	612	5.6	13.7	197	296	82
  051	622	12.9	8.3	119	246	150
  139	628	8.5	7.8	145	166	83
  150	628	8.6	14.5	295**	315	183
  043	635	5.9	13.7	239	272	189
  096	651	17.4	9.7	326**
  073	657	7.0	9.5	186	283	78
  127	665	5.8	8.1	166	344**	147
  121	677	10.2	9.5	226	346**	173
  034	694	15.9	12.1	406**	592**	584**
  124	697	9.7	11.0	63	179	60
  123	702	10.4	10.6	186	148	96
  113	705	7.6	8.4	107	534**	92
  071	709	10.6	11.3	207	584**	141
  076	722	8.1	10.5	271	489**	138
  044	724	7.3	12.1	212	683**	217
  040	731	15.1	7.4	205	149	136
  062	741	4.4	18.7**	153	856**	416**
  025	741	10.0	12.2	224	344**	121
  119	755	5.9	10.1	187	377**	61
  075	757	10.0	24.7**	246	345**	276**
  098	759	13.8	13.9	380**	239	156
  134	769	7.5	10.4	125	131	81
  087	773	25.0	10.1	181	285	135
  142	788	4.6	12.1	166	273	129
  064	792	15.4	8.6	218	299	139
  094	793	16.6	10.0	186	179	173
  022	808	8.8	14.4	184	271	161
  112	812	12.0	9.2	181	184	108
  125	817	14.4	11.0	158	242	72
  106	862	5.3	9.2	94	300	95
  146	890	13.9	11.9	135
  058	897	5.3	18.5**	154	460**	80
  063	943	17.8	19.7**	277**	642**	306**
  095	960	25.3	10.7	135	181	111
  152	963	9.4	8.8	198
  130	971	15.9	13.5	106	307	84
  059	1063	9.4	9.7	129	378**	54
  105	1109	11.0	6.1	87	155	64
  136	1163	6.0	13.1	250	565**	122
  065	1251	14.5	10.7	88	147	88
  029	1490	22.2	9.7	129	111	105
  144	1536	7.0	17.7**	216	694**	418**
  068	1809	12.7	10.4	59	128	39

• TABLE 3
  SERUM METABOLITE & VITAMIN LEVELS IN A GERIATRIC
  NURSING HOME POPULATION

  Patient	B12	Folate	Homocysteine	Methylmalonic Acid

  NH170	8**	14.0	34.8**	3365**
  NH129	40**	7.4	40.9**	6245**
  NH156	44**	22.4	17.4**	1130**
  NH139	56**	9.7	20.9**	1180**
  NH132	67**	7.6	92.4**	12641**
  NH176	129**	9.2	20.3**	1156**
  NH196	136**	6.2	41.0**	1077**
  NH109	139**	9.8	20.9**	1294**
  NH203	146**	4.3	12.2	437**
  NH141	161**	13.4	12.2	223
  NH178	172**	8.2	5.9	141
  NH103	189**	5.5	13.1	362**
  NH181	196**	6.3	14.7	296**
  NH160	206	11.9	12.5	640**
  NH197	221	24.0	10.5	654**
  NH073	222	3.6**	19.8**	490**
  NH110	227	5.5	13.7	1297**
  NH010	228	4.0	21.1**	413**
  NH012	234	8.7	16.0	596**
  NH037	236	11.5	22.5**	11299**
  NH114	238	12.8	13.2	442**
  NH211	240	6.0	14.1	166
  NH075	250	9.3	12.1	170
  NH172	255	7.2	14.4	552**
  NH148	259	5.7	19.2**	317**
  NH138	264	9.2	16.7**	340**
  NH150	264	4.0	13.7	98
  NH099	272	5.5	12.5	125
  NH124	275	6.9	11.5	87
  NH179	301	7.6	7.1	143
  NH135	302	6.5	23.4**	397**
  NH087	304	7.8	10.8	327**
  NH180	304	5.8	10.5	237
  NH209	306	7.6	11.9	105
  NH107	310	3.3**	8.6	148
  NH081	320	4.3	23.6**	470**
  NH068	324	7.9	13.4	243
  NH183	325	7.7	11.1	144
  NH033	330	13.8	7.7	149
  NH161	333	8.5	11.3	385**
  NH192	337	10.7	9.5	209
  NH136	340	6.7	18.2**	409**
  NH191	342	20.2	13.4	271
  NH137	343	4.0	15.6	183
  NH182	346	8.2	14.4	448**
  NH020	347	8.4	10.4	149
  NH165	351	18.5	11.8	425**
  NH095	352	8.5	14.5	366**
  NH194	361	4.3	20.3**	305**
  NH106	362	4.8	12.9	298**
  NH060	367	4.7	16.4**	71
  NH009	368	5.1	15.9	325**
  NH071	382	4.9	12.9	330**
  NH080	390	6.1	15.0	171
  NH013	407	6.7	12.4	310**
  NH126	409	9.2	17.4**	137
  NH030	411	11.2	10.4	844**
  NH210	413	8.6	11.9	210
  NH158	414	5.7	16.2	508**
  NH027	416	10.2	15.5	769**
  NH003	424	16.5	9.5	167
  NH187	429	4.7	8.8	439**
  NH022	430	10.5	14.0	214
  NH082	436	10.6	17.7**	340**
  NH162	438	6.1	19.2**	180
  NH021	439	5.3	15.1	191
  NH056	447	11.7	10.9	184
  NH119	448	3.2**	14.1	241
  NH120	448	5.6	12.0	138
  NH186	450	4.7	23.1**	213
  NH064	451	6.9	10.6	237
  NH057	453	14.6	10.4	282**
  NH131	454	8.1	16.2	258
  NH059	462	6.0	9.1	147
  NH202	465	3.3**	17.0**	393**
  NH134	475	15.3	11.6	321**
  NH083	475	7.4	10.6	178
  NH199	479	15.1	10.4	141
  NH042	482	6.0	15.0	141
  NH200	491	13.6	9.8	154
  NH213	497	8.1	10.0	92
  NH143	500	5.2	22.1**	175
  NH031	502	6.4	16.1	151
  NH188	504	12.5	15.1	1461**
  NH171	504	10.7	12.9	344**
  NH008	505	4.6	9.9	185
  NH102	506	16.6	9.1	236
  NH145	512	7.7	22.2**	161
  NH093	514	5.1	17.7**	185
  NH118	524	25.0	10.1	314**
  NH185	524	8.7	12.1	84
  NH111	527	5.1	18.4**	250
  NH149	530	12.6	18.2**	531**
  NH011	534	8.1	12.5	654**
  NH128	540	4.3	11.6	120
  NH035	547	7.5	9.8	193
  NH005	551	17.7	5.0	365**
  NH212	552	11.9	12.1	202
  NH007	554	6.4	26.1**	646**
  NH086	554	9.5	5.1	127
  NH069	555	22.7	6.8	134
  NH121	555	8.2	10.0	112
  NH117	571	6.6	9.7	351**
  NH055	581	14.8	9.1	265
  NH025	581	5.2	15.3	181
  NH104	583	3.9	14.6	1699**
  NH173	583	11.2	10.6	160
  NH177	584	6.2	5.7	111
  NH207	586	8.5	16.4**	243
  NH070	591	5.4	12.0	168
  NH038	592	8.0	8.8	230
  NH049	599	10.7	21.7**	238
  NH062	606	4.5	7.7	96
  NH153	608	7.7	13.6	221
  NH206	611	6.6	16.4**	400**
  NH018	614	6.3	10.9	123
  NH163	616	5.0	9.6	132
  NH189	619	7.6	12.0	158
  NH045	620	21.0	12.4	265
  NH074	621	10.2	9.2	172
  NH054	623	8.0	9.8	121
  NH152	625	8.2	7.8	206
  NH140	637	21.7	13.6	300**
  NH050	642	16.3	13.5	275**
  NH089	644	7.7	16.7**	444**
  NH036	649	7.9	10.7	68
  NH097	651	6.6	13.4	426**
  NH016	656	4.1	61.0**	356**
  NH053	657	14.2	10.6	320**
  NH066	658	7.7	11.4	228
  NH051	659	4.0	10.7	216
  NH108	671	5.8	24.0**	823**
  NH058	673	6.0	11.2	392**
  NH028	675	22.3	9.1	105
  NH204	678	4.7	10.2	148
  NH169	679	6.9	19.2**	267
  NH032	681	12.7	5.9	99
  NH065	682	11.0	13.5	176
  NH061	683	13.4	9.6	190
  NH116	685	9.0	7.5	244
  NH015	699	6.8	16.8**	236
  NH157	711	10.0	12.8	198
  NH155	715	10.0	17.6**	308**
  NH034	715	7.9	11.4	179
  NH040	717	10.5	15.7	256
  NH105	718	6.0	13.2	308**
  NH048	719	8.0	10.8	207
  NH084	720	6.8	9.4	169
  NH115	724	16.3	9.4	161
  NH205	734	8.5	13.3	232
  NH113	738	11.7	10.3	171
  NH154	738	13.7	9.6	123
  NH167	741	17.0	6.6	129
  NH190	752	5.2	14.1	254
  NH067	760	22.5	9.5	232
  NH014	767	8.9	7.3	100
  NH072	768	8.3	6.9	131
  NH133	772	8.8	20.4**	219
  NH122	778	6.0	10.4	108
  NH076	781	12.1	14.9	282**
  NH147	785	7.5	24.5**	411**
  NH026	786	9.7	8.3	146
  NH151	789	24.4	11.1	182
  NH198	797	10.9	10.7	158
  NH088	801	6.4	18.3**	184
  NH004	806	11.3	8.8	96
  NH024	818	5.1	14.1	219
  NH100	826	16.4	10.5	103
  NH078	831	7.2	10.3	266
  NH052	844	19.6	8.0	193
  NH142	848	18.6	12.1	398**
  NH002	862	9.4	11.3	212
  NH091	891	4.9	12.6	169
  NH127	897	22.0	8.4	132
  NH096	901	9.3	5.2	104
  NH201	910	25.0	15.7	424**
  NH184	941	21.5	10.8	170
  NH208	945	20.2	9.8	111
  NH130	968	22.4	10.4	339**
  NH164	989	8.0	16.8**	102
  NH077	1006	15.1	9.2	188
  NH017	1015	11.9	9.5	175
  NH029	1053	18.6	11.4	161
  NH023	1055	9.3	9.7	193
  NH047	1079	6.4	11.4	106
  NH043	1082	14.5	13.9	144
  NH195	1088	36.9	12.2	150
  NH193	1092	8.2	15.7	225
  NH046	1093	9.2	18.8**	186
  NH101	1108	3.9	8.1	139
  NH098	1117	11.3	12.5	88
  NH168	1124	25.2	15.0	203
  NH006	1126	6.9	8.1	159
  NH144	1135	8.0	21.9**	262
  NH044	1159	26.8	10.2	109
  NH175	1162	7.8	12.0	210
  NH146	1179	9.8	10.1	129
  NH112	1238	10.3	15.0	347**
  NH001	1304	13.1	6.9	142
  NH166	1337	13.4	8.3	67
  NH079	1346	18.0	12.0	248
  NH041	1528	20.7	8.2	155
  NH063	1559	15.0	7.0	66
  NH159	1566	6.6	15.5	451**
  NH125	1703	8.2	20.6**	153
  NH094	1768	15.9	8.4	182
  NH123	2028	10.2	16.8**	206
  NH174	2106	13.3	12.8	280**
  NH039	2227	23.8	8.9	119
  NH019	2297	11.1	15.5	177
  NH092	2360	5.7	9.8	131
  NH085	3141	22.0	26.9**	1947**

• TABLE 4
  SERUM METABOLITE & VITAMIN LEVELS IN A GERIATRIC
  POPULATION

  	Patient	B12	Folate	Homocysteine	MMA

  	495	77**	10.0	65.4**	3145**
  	484	84**	10.0	77.5**	6820**
  	522	100**	3.6**	15.5	967**
  	455	115**	1.9**	21.8**	170
  	493	135**	4.4	16.9**	421**
  	528	145**	3.9	38.3**	729**
  	510	155**	4.6	14.1	804**
  	502	155**	2.1**	16.9**	347**
  	412	160**	18.5**	33.8**	1301**
  	409	160**	4.8	16.8**	164
  	470	165**	9.2	19.9**	1468**
  	460	165**	6.8	11.5	142
  	437	170**	4.9	16.5**	813
  	439	170**	1.2**	21.3**	502**
  	525	175**	11.5	15.3	1058**
  	442	175**	4.2	17.5**	328**
  	456	180**	7.3	11.1	206
  	450	180**	5.0	11.8	196
  	477	185**	3.4**	31.4**	369**
  	508	190**	4.1	19.5**	335**
  	423	190**	2.5**	19.0**	329**
  	462	190**	3.8	11.6	276**
  	523	190**	5.6	16.8**	207
  	482	190**	2.9**	25.1**	179
  	459	190**	5.3	19.6**	167
  	543	195**	4.3	13.5	470**
  	520	195**	1.7**	22.2**	309**
  	431	195**	7.2	13.5	251
  	513	200	5.0	25.0**	1184**
  	534	200	4.9	32.6**	1080**
  	515	200	4.9	17.3**	478**
  	531	200	5.1	26.8**	466**
  	516	200	3.6**	17.8**	279**
  	526	200	1.6**	23.5**	171
  	471	205	5.7	22.0**	542**
  	413	205	2.6**	20.4**	304**
  	497	205	3.3**	19.4**	258
  	539	205	4.1	15.4	247
  	544	205	12.5	11.7	233
  	540	205	4.0	17.1**	185
  	517	205	2.2**	15.0	151
  	496	210	3.7**	15.2	1103**
  	488	210	16.5	21.8**	600**
  	416	215	12.5	10.0	197
  	434	220	7.1	24.8**	439**
  	545	220	11.5	14.4	407**
  	547	220	5.3	17.5**	396**
  	408	220	3.2**	16.4**	357**
  	449	220	3.7**	13.7	272**
  	507	220	8.5	10.0	179
  	458	225	10.5	21.1**	964**
  	491	225	7.2	16.0	472**
  	529	230	2.0**	61.1	1172**
  	415	230	3.2**	28.9**	377**
  	453	230	3.6**	19.8**	336**
  	448	230	5.2	13.1	319**
  	498	230	5.9	20.1**	255
  	533	230	5.7	11.7	151
  	466	235	35.0	12.1	617**
  	537	235	5.7	10.7	394**
  	483	235	8.6	16.6**	344**
  	512	235	3.9	12.5	190
  	452	240	4.7	26.5**	1068**
  	454	240	5.2	11.9	201
  	535	240	4.4	15.3	195
  	421	245	10.5	12.5	464**
  	469	245	6.2	20.0**	448**
  	474	245	7.3	10.3	327**
  	486	245	9.2	12.6	156
  	536	250	22.5	20.3**	1068**
  	475	250	5.6	23.0	456**
  	511	250	2.7**	23.1**	398**
  	465	250	4.1	23.1**	323**
  	506	250	5.2	11.5	252
  	417	250	5.5	25.2**	241
  	524	1250	2.5**	14.4	212
  	411	250	9.9	11.5	200
  	492	250	5.2	10.7	182
  	548	250	2.9**	12.4	179
  	441	250	4.5	8.5	147
  	480	255	4.8	16.9**	558**
  	532	255	7.0	14.8	419**
  	464	255	11.5	12.9	400**
  	494	255	6.2	12.1	293**
  	106	255	4.5	11.7	203
  	546	260	5.5	14.7	662**
  	541	260	5.4	30.8**	426**
  	420	260	5.3	13.6	347**
  	500	260	6.7	14.0	330**
  	538	260	9.3	17.3**	298**
  	457	260	2.9**	12.6	286**
  	472	260	8.3	13.8	278**
  	424	260	8.3	10.1	242
  	433	260	6.8	10.5	197
  	425	265	7.3	14.7	724**
  	468	265	3.8	16.7**	289**
  	435	265	7.4	14.0	150
  	499	265	2.2**	12.4	131
  	432	270	4.3	28.3**	432**
  	521	270	3.7**	15.3	349**
  	549	270	4.21	12.4	343**
  	518	270	10.0	10.1	276**
  	418	270	26.0	9.4	213
  	419	270	6.5	12.5	212
  	428	270	4.2	18.7**	189
  	443	270	8.8	12.0	187
  	446	270	11.0	8.1	157
  	461	275	7.6	15.1	663**
  	440	275	4.9	12.9	248
  	436	275	6.3	30.1**	233
  	530	275	7.4	13.6	231
  	438	275	4.6	8.5	221
  	527	275	7.5	10.5	219
  	444	275	4.0	12.2	180
  	429	280	5.3	15.3	463**
  	503	280	4.4	25.7**	421**
  	485	280	3.5**	15.6	381**
  	410	280	14.5	10.0	201
  	487	280	3.9	10.5	166
  	430	280	9.2	8.8	161
  	519	285	3.9	22.2**	919**
  	476	285	10.5	12.8	339**
  	509	285	5.4	13.0	331**
  	501	285	5.5	12.4	252
  	542	285	6.9	15.5	242
  	445	285	7.2	14.9	237
  	427	285	4.0	17.1**	233
  	490	290	4.7	13.9	203
  	451	290	2.1**	20.0**	226
  	414	290	7.0	9.7	117
  	467	290	4.1	6.5	68
  	463	295	5.8	12.3	296**
  	473	295	7.5	14.4	290**
  	505	295	4.1	12.4	257
  	198	300	11.5	10.9	323**
  	195	300	9.8	12.2	216
  	207	305	7.7	13.2	330**
  	67	305	8.6	15.4	312**
  	50	305	9.0	11.6	235
  	70	305	12.5	12.7	228
  	113	305	5.6	13.5	201
  	39	305	6.9	19.7**	170
  	3	305	4.2	11.5	135
  	325	305	14.5	9.4	94
  	368	310	4.7	15.9	371**
  	322	310	7.8	15.3	362**
  	295	310	7.2	13.8	305**
  	347	310	5.8	16.5**	266
  	313	310	6.1	16.5**	219
  	355	310	5.5	15.4	138
  	291	310	4.5	15.2	125
  	478	315	23.0	17.7**	857**
  	53	315	5.8	12.1	505**
  	240	315	6.7	12.3	394**
  	14	315	9.6	14.2	331**
  	137	315	7.8	24.3**	306**
  	254	315	8.7	17.0**	285**
  	109	315	3.7**	16.5**	263
  	252	315	5.2	10.1	241
  	186	315	4.1	15.4	238
  	183	315	5.5	10.7	195
  	390	315	6.9	10.0	188
  	267	315	2.2**	12.0	124
  	310	320	12.0	13.8	395**
  	31	320	17.0	12.9	334**
  	88	320	4.8	13.8	217
  	403	320	9.6	11.3	162
  	60	320	6.2	11.4	155
  	315	320	6.4	9.9	136
  	175	325	6.3	17.8**	486**
  	317	325	22.0	14.0	294**
  	18	325	6.3	11.1	241
  	247	325	13.5	13.2	231
  	223	325	9.2	12.6	203
  	132	325	3.7**	15.4	184
  	168	325	4.3	10.2	174
  	238	325	5.5	9.9	166
  	117	325	5.2	15.0	154
  	404	330	2.5**	33.1**	1085**
  	138	330	4.8	11.3	360**
  	316	330	3.6**	10.2	272**
  	61	330	5.1	12.5	242
  	333	330	34.0	9.2	235
  	16	330	4.6	13.3	211
  	276	330	5.7	11.9	200
  	391	330	4.1	8.4	184
  	362	330	9.2	11.7	178
  	1	330	9.9	8.9	170
  	379	335	16.0	12.1	471**
  	147	335	9.0	9.7	427**
  	89	335	8.0	15.3	385**
  	211	335	5.0	12.2	374**
  	45	335	5.9	16.3**	250
  	47	335	5.0	13.6	249
  	402	335	4.7	13.5	230
  	314	335	7.6	9.7	203
  	150	335	4.8	11.2	119
  	120	340	1.9**	21.0**	775**
  	284	340	7.2	25.6**	439**
  	230	340	14.0	11.4	419**
  	149	340	8.8	18.9**	337**
  	269	340	3.9	16.2	302**
  	197	340	10.5	12.8	233
  	19	340	9.6	11.0	232
  	422	340	3.1**	14.4	188
  	196	340	11.5	8.9	169
  	40	345	8.7	14.6	610**
  	244	345	8.6	15.8	461**
  	287	345	5.7	18.1**	427**
  	100	345	8.3	14.8	403**
  	383	345	4.3	27.2**	284**
  	62	345	19.5	9.6	250
  	350	345	8.0	10.0	249
  	65	345	8.0	10.2	247
  	307	345	16.5	11.6	208
  	69	345	17.0	9.9	197
  	328	345	7.5	8.9	192
  	43	345	6.0	13.2	191
  	222	345	6.1	9.2	175
  	306	345	4.3	17.2**	160
  	154	345	7.1	10.2	148
  	94	350	4.8	16.1	302**
  	201	350	6.1	9.9	200
  	13	350	5.1	10.9	193
  	236	355	7.2	14.8	309**
  	191	355	5.8	15.3	257
  	481	355	5.2	17.1**	134
  	92	360	4.2	25.2**	321**
  	324	360	3.8	16.6**	264
  	87	360	3.3**	13.3	200
  	46	360	5.4	11.1	179
  	289	360	9.5	7.9	129
  	392	360	5.1	10.3	125
  	320	365	6.4	17.3**	240
  	134	365	13.5	11.8	238
  	239	365	7.7	13.2	236
  	326	365	6.0	10.9	180
  	364	365	4.1	13.9	154
  	218	365	7.5	11.2	126
  	216	365	6.2	12.2	119
  	248	365	5.7	13.3	117
  	375	370	4.1	20.7**	532**
  	288	370	6.4	18.8**	436**
  	161	370	6.3	11.2	340**
  	244	370	19.5	9.8	286**
  	330	370	18.0	12.2	228
  	334	370	12.5	8.7	172
  	275	370	6.9	12.7	162
  	54	375	7.3	10.1	583**
  	185	375	9.3	10.5	386**
  	52	375	8.1	15.5	291**
  	366	375	5.0	12.5	280**
  	93	375	3.3**	16.2	248
  	151	375	2.9**	12.3	235
  	85	375	6.7	14.8	217
  	294	375	7.0	12.2	184
  	361	375	7.9	10.7	179
  	318	375	5.5	13.7	160
  	386	375	7.6	10.4	153
  	304	375	9.1	9.4	132
  	228	380	7.7	17.1**	320**
  	110	380	4.0	7.2	135
  	204	380	5.7	10.6	91
  	348	385	2.3**	17.4**	368**
  	146	385	11.5	12.5	253
  	260	385	5.5	13.7	211
  	136	385	3.6**	19.8**	205
  	338	385	5.0	16.2	180
  	376	385	3.6**	13.7	154
  	194	385	12.5	7.9	153
  	504	385	38.0	9.5	138
  	160	390	8.1	24.7**	475**
  	354	390	11.5	12.8	212
  	25	390	5.1	11.3	205
  	387	390	8.7	8.4	162
  	86	390	21.0	12.6	133
  	133	390	3.9	11.3	113
  	331	395	12.0	20.1**	638**
  	130	395	10.5	10.8	256
  	82	395	2.8**	9.8	236
  	119	395	12.5	16.3**	209
  	380	395	10.5	14.3	159
  	373	395	5.5	11.6	152
  	256	395	10.5	9.9	149
  	384	395	7.3	14.7	116
  	105	400	19.0	10.5	322**
  	251	400	4.8	14.9	289**
  	352	400	11.5	9.6	181
  	279	400	4.5	11.7	170
  	339	400	7.4	13.6	168
  	381	405	6.7	12.4	294**
  	285	405	7.0	14.2	281**
  	340	405	3.6**	19.6**	275**
  	51	405	6.5	14.3	233
  	33	405	6.5	9.6	207
  	268	405	3.3**	14.9	205
  	73	405	5.2	13.1	172
  	17	410	7.5	16.2	473**
  	286	410	4.7	18.8**	415**
  	140	410	5.9	21.7**	302**
  	116	410	6.8	14.5	218
  	396	410	5.6	16.1	190
  	356	410	1.9**	27.6**	149
  	237	410	3.6**	16.6**	122
  	112	410	5.5	8.9	107
  	259	410	4.7	11.6	99
  	176	415	5.2	21.9**	453**
  	193	415	10.5	11.3	163
  	323	415	6.1	9.6	163
  	202	415	11.5	9.4	150
  	398	415	8.0	12.6	134
  	321	420	5.2	10.7	383**
  	142	420	29.0	8.3	234
  	327	420	3.2**	14.6	203
  	342	420	7.3	9.4	156
  	170	420	20.5	10.3	142
  	345	420	29.5	13.2	136
  	302	420	8.6	8.8	128
  	115	425	6.3	22.2**	628**
  	97	425	12.5	19.8**	313**
  	246	425	8.7	15.1	241
  	72	425	10.5	13.5	241
  	365	425	6.7	16.7**	237
  	139	425	12.5	10.4	224
  	143	425	8.1	13.5	216
  	426	425	19.5	14.5	201
  	303	425	3.0**	14.5	154
  	388	425	6.2	12.3	135
  	127	425	6.7	8.4	100
  	262	430	10.0	12.1	323**
  	270	430	4.8	12.9	293**
  	514	430	4.3	12.9	197
  	341	430	3.5**	19.9**	190
  	278	430	5.2	10.8	182
  	370	430	11.0	15.3	174
  	55	430	7.6	11.0	162
  	274	430	5.0	8.2	131
  	367	430	17.5	8.0	126
  	98	430	13.5	12.8	125
  	337	435	13.5	14.1	395**
  	309	435	8.7	12.9	349**
  	305	435	17.5	15.4	187
  	144	435	25.0	8.9	167
  	34	435	8.6	7.6	157
  	234	435	9.7	9.2	116
  	123	440	9.6	12.2	622**
  	200	440	4.8	12.4	257
  	250	440	7.5	12.9	248
  	107	440	6.3	14.7	183
  	300	440	6.5	7.9	123
  	374	445	5.4	14.0	247
  	372	445	11.0	11.0	181
  	36	445	4.0	10.0	181
  	271	445	7.2	10.4	124
  	242	445	15.5	9.6	112
  	264	445	6.0	10.7	100
  	172	450	11.5	14.9	607**
  	32	450	11.5	13.6	362**
  	346	450	13.5	15.8	330**
  	41	450	8.5	11.4	194
  	95	450	5.1	12.5	182
  	357	455	6.3	14.4	296**
  	319	455	17.0	10.2	147
  	308	455	15.0	9.8	131
  	235	455	23.0	9.0	114
  	349	455	9.2	8.3	82
  	178	460	5.6	20.6**	473**
  	312	460	4.7	14.4	197
  	79	460	5.0	10.4	173
  	131	460	18.0	10.2	162
  	243	460	2.6**	11.6	160
  	261	465	7.7	10.6	252
  	378	465	5.4	13.2	221
  	49	465	47.0	10.8	179
  	226	465	7.7	10.2	173
  	377	465	5.6	8.5	143
  	253	465	10.0	7.0	138
  	76	470	12.5	14.8	304**
  	203	470	15.0	7.6	233
  	296	470	23.5	11.0	161
  	382	470	5.3	11.1	109
  	6	475	10.5	12.5	232
  	75	475	4.5	8.1	150
  	332	475	9.4	10.0	144
  	290	475	14.0	9.1	143
  	128	475	5.9	9.3	133
  	124	475	6.0	13.5	111
  	177	475	8.8	9.1	106
  	126	480	11.0	11.0	212
  	283	480	5.2	10.6	175
  	209	480	10.5	10.5	175
  	293	480	6.8	15.5	135
  	121	485	4.7	20.0**	345**
  	282	485	12.0	10.9	236
  	71	485	13.5	8.1	168
  	385	485	9.0	14.1	128
  	190	495	9.9	10.4	410**
  	210	495	8.6	12.0	243
  	155	495	5.9	10.4	219
  	336	495	13.5	9.9	135
  	280	500	8.7	14.5	334**
  	96	500	4.7	10.8	237
  	145	500	5.9	17.5**	233
  	199	500	4.2	13.8	199
  	489	500	11.5	9.7	198
  	217	500	6.4	9.6	166
  	90	500	7.5	8.5	106
  	164	510	5.2	23.8**	408**
  	343	510	4.5	13.7	284**
  	42	510	4.9	7.4	233
  	351	510	8.5	11.0	207
  	299	510	12.0	8.0	104
  	99	520	10.5	25.8**	322**
  	114	520	30.0	10.9	220
  	369	520	29.0	16.7**	206
  	37	520	10.5	8.6	191
  	215	520	6.7	16.8**	151
  	401	520	7.5	12.6	148
  	229	520	7.9	11.0	116
  	135	520	3.2**	8.3	88
  	81	530	6.8	14.8	372**
  	91	530	14.5	10.6	228
  	167	530	23.5	9.2	176
  	181	530	5.5	9.3	171
  	56	530	20.0	8.3	163
  	5	530	13.5	8.1	159
  	180	540	12.0	9.0	216
  	311	540	4.1	13.3	214
  	389	540	3.9	13.9	169
  	125	540	5.5	13.0	159
  	35	540	22.5	11.0	123
  	104	550	10.5	16.5**	544**
  	393	550	4.9	11.9	339**
  	394	550	23.0	14.0	278**
  	292	550	6.9	16.2	263
  	163	550	6.7	14.3	219
  	66	550	10.5	11.6	206
  	29	550	17.5	9.6	191
  	227	550	7.9	11.7	154
  	38	550	7.5	11.9	152
  	241	550	10.5	9.8	100
  	102	550	9.7	8.6	91
  	77	560	24.0	14.8	554**
  	162	560	10.5	11.8	275**
  	273	560	8.7	9.4	180
  	80	560	6.3	11.2	108
  	255	560	8.8	9.9	93
  	122	570	66.0	13.8	304**
  	208	570	34.0	10.2	255
  	23	570	21.5	8.3	241
  	447	570	25.0	10.0	164
  	225	570	5.7	12.2	154
  	174	570	7.1	11.0	127
  	11	570	19.0	8.9	113
  	165	580	10.5	14.8	226
  	182	580	8.9	8.2	189
  	245	590	15.5	10.0	262
  	83	590	17.5	8.3	199
  	166	590	11.5	9.4	188
  	158	590	7.3	10.7	166
  	187	590	4.5	11.0	146
  	156	590	23.5	11.3	112
  	231	600	9.5	9.0	192
  	78	600	11.5	9.4	151
  	329	610	15.0	7.3	312**
  	57	610	16.0	11.9	286**
  	7	610	12.0	10.4	195
  	277	610	9.5	7.8	153
  	108	620	13.5	8.4	191
  	205	620	18.0	7.5	145
  	263	620	9.8	10.2	101
  	9	630	4.9	11.4	300**
  	111	630	8.3	11.1	276**
  	68	630	11.5	8.9	143
  	399	630	14.0	11.0	90
  	266	640	5.1	15.7	364**
  	12	640	24.5	9.0	233
  	152	640	8.1	10.0	209
  	405	640	7.0	12.8	186
  	27	640	22.5	8.4	136
  	258	640	8.3	11.2	120
  	249	640	8.7	9.1	81
  	297	650	16.0	10.0	279**
  	192	650	4.9	14.9	213
  	257	650	3.3**	16.3**	208
  	184	650	12.5	9.9	193
  	58	650	18.5	10.7	172
  	301	650	16.0	15.5	162
  	397	650	12.5	8.4	146
  	272	650	11.0	7.4	120
  	153	650	7.1	13.1	116
  	406	650	6.6	5.8	81
  	10	660	9.0	7.6	154
  	26	660	22.0	8.3	132
  	265	670	3.9	19.3**	509**
  	359	670	21.0	8.3	269
  	48	670	32.0	9.9	262
  	335	670	11.5	8.1	121
  	189	680	6.6	17.9**	358**
  	220	680	15.5	10.9	115
  	15	690	13.5	13.4	159
  	44	700	20.0	12.7	244
  	21	700	13.5	10.2	129
  	74	700	15.0	7.1	65
  	4	710	29.0	8.5	266
  	353	710	11.5	11.4	206
  	281	710	10.5	9.6	185
  	2	710	8.0	8.5	109
  	212	740	20.0	11.1	250
  	8	740	12.0	11.5	216
  	206	750	12.5	8.3	116
  	101	770	14.5	12.7	372**
  	344	770	32.0	11.7	297**
  	20	770	35.0	10.1	245
  	407	770	10.5	12.0	110
  	360	780	2.7**	20.9**	157
  	232	790	15.5	10.1	151
  	141	790	12.5	9.5	74
  	129	800	8.7	11.7	211
  	188	800	15.0	12.3	174
  	400	800	12.5	10.3	156
  	24	810	23.0	7.5	194
  	173	830	35.0	11.4	243
  	214	830	21.5	12.0	187
  	63	830	13.8	8.8	185
  	148	830	45.0	7.1	146
  	84	830	23.5	7.0	136
  	179	830	16.5	6.6	96
  	171	840	23.5	11.2	195
  	28	870	5.8	15.9	197
  	233	870	7.9	12.7	169
  	221	870	40.0	7.0	126
  	371	880	20.0	8.5	152
  	213	890	10.5	18.0**	231
  	358	900	21.0	8.3	149
  	298	910	15.5	10.2	221
  	118	910	100.0	9.7	170
  	479	950	11.5	12.1	188
  	30	950	6.2	10.5	170
  	159	1000	9.5	8.7	281**
  	219	1050	37.0	14.3	313**
  	103	1050	12.5	10.3	154
  	59	1150	17.5	7.3	180
  	157	1250	12.0	14.0	206
  	363	1350	28.0	10.4	190
  	22	1400	13.5	10.4	233
  	64	1400	31.0	9.7	149
  	169	1450	15.0	9.5	150

Claims (32)

1. A method of preventing vascular disease in a human, comprising periodically administering orally a single formulation having between 0.3 and 10 mg vitamin B₁₂ and 0.1 and 0.4 mg folic acid.

2. The method of claim 1 wherein the formulation includes approximately 2 mg vitamin B₁₂ and 0.4 mg folic acid.

3. The method of claim 1 wherein said vascular disease is cardiovascular disease.

4. The method of claim 1 wherein said vascular disease is stroke.

5. The method of claim 1 wherein said vascular disease is peripheral vascular disease.

6. The method of claim 1 wherein said vascular disease is associated with renal disease.

7. A method of preventing vascular disease in a human, comprising periodically administering orally a single formulation having between 0.3 and 10 mg vitamin B_12, and 0.1 and 0.4 mg folic acid, and 5 and 75 mg vitamin B6.

8. The method of claim 7 wherein the formulation includes approximately 2 mg vitamin B12 and 0.4 mg folic acid and 25 mg vitamin B6.

9. The method of claim 7 wherein said vascular disease is cardiovascular disease.

10. The method of claim 7 wherein said vascular disease is stroke.

11. The method of claim 7 wherein said vascular disease is peripheral vascular disease.

12. The method of claim 7 wherein said vascular disease is associated with renal disease.

13. A method of preventing vascular disease in a human, comprising periodically administering orally a single formulation having between 0.3 and 10 mg vitamin B12 and 0.4 and 10.0 mg folic acid.

14. The method of claim 13 wherein the formulation includes approximately 2 mg vitamin B12 and 1.0 mg folic acid.

15. The method of claim 13 wherein the formulation includes approximately 2 mg vitamin B12 and 2.0 mg folic acid.

16. The method of claim 13 wherein the formulation includes approximately 2 mg vitamin B12 and 2.5 mg folic acid.

17. The method of claim 13 wherein the formulation includes approximately 1 mg vitamin B12 and 2.0 mg folic acid.

18. The method of claim 13 wherein the formulation includes approximately 1 mg vitamin B12 and 2.5 mg folic acid.

19. The method of claim 13 wherein said vascular disease is cardiovascular disease.

20. The method of claim 13 wherein said vascular disease is stroke.

21. The method of claim 13 wherein said vascular disease is peripheral vascular disease.

22. The method of claim 13 wherein said vascular disease is associated with renal disease.

23. A method of preventing vascular disease in a human, comprising periodically administering orally a single formulation having between 0.3 and 10 mg vitamin B12, and 0.4 and 10 mg folic acid, and 5 and 75 mg vitamin B6.

24. The method of claim 23 wherein the formulation includes approximately 2 mg vitamin B12 and 1.0 mg folic acid and 25 mg vitamin B6.

25. The method of claim 23 wherein the formulation includes approximately 2 mg vitamin B12 and 2.0 mg folic acid and 25 mg vitamin B6.

26. The method of claim 23 wherein the formulation includes approximately 2 mg vitamin B12 and 2.5 mg folic acid and 25 mg vitamin B6.

27. The method of claim 23 wherein the formulation includes approximately 1 mg B12 and 2.0 mg folic acid and 25 mg vitamin B6.

28. The method of claim 23 wherein the formulation includes approximately 1 mg B12 and 2.5 mg folic acid and 25 mg vitamin B6.

29. The method of claim 23 wherein said vascular disease is cardiovascular disease.

30. The method of claim 23 wherein said vascular disease is stroke.

31. The method of claim 23 wherein said vascular disease is peripheral vascular disease.

32. The method of claim 23 wherein said vascular disease is associated with renal disease.

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