INJECTABLE NUTRITIONAL SUPPLEMENT

Abstract

An aqueous injectable nutritional supplement composition for livestock, comprising: an EDTA complex of one or more trace elements; vitamin B.sub.12; and a water-soluble liquid selected from glycols, glycol ethers and mixtures thereof.

Claims

1. An aqueous injectable nutritional supplement composition for livestock, comprising: an EDTA complex of one or more trace elements; vitamin B.sub.12; and a water-soluble liquid selected from glycols, glycol ethers and mixtures thereof, and wherein the trace elements are in an amount of 1 g to 100 g per litre of aqueous injectable composition, wherein the one or more trace elements comprise at least one of zinc, copper, manganese, selenium and chromium.

2. (canceled)

3. The aqueous injectable nutritional supplement of claim 1, wherein the one or more trace elements comprise at least one of zinc and copper.

4. The aqueous injectable nutritional supplement of claim 1, wherein the trace elements are present in an amount of 20 g to 100 g per litre.

5. The aqueous injectable nutritional supplement of claim 1, wherein the composition trace elements include 10 g to 60 g of zinc per litre, 5 g to 30 g of copper per litre or 10 g to 60 g of zinc per litre and 5 g to 30 g of copper per litre.

6. The aqueous injectable nutritional supplement composition of claim 1, wherein the water-soluble liquid is selected from the group consisting of propylene glycol and glycol ethers of molecular weight no more than 300 and mixtures thereof.

7. The aqueous injectable nutritional supplement composition of claim 1, wherein the water-soluble liquid is selected from propylene glycol polyethylene glycol of molecular weight no more than 300, dipropylene glycol monomethyl ether and mixtures thereof.

8. The aqueous injectable nutritional supplement composition of claim 1, wherein the trace elements comprise at least one further trace element selected from manganese, selenium and chromium.

9. The aqueous injectable nutritional supplement composition of claim 1, wherein the trace elements comprise zinc, copper and selenium.

10. The aqueous injectable nutritional supplement composition of claim 1, wherein the vitamin B.sub.12 comprises at least one of cyanocobalamin, hydroxocobalamin, hydroxocobalamin acetate, methylcobalamin and aquacobalamin and mixtures thereof in an amount of 0.5 g to 8 g per litre of aqueous injectable composition.

11. The aqueous injectable composition of claim 10, wherein the amount of vitamin B.sub.12 is an amount of 1 g to 8 g.

12. The aqueous injectable nutritional supplement composition of claim 1, wherein the water-soluble liquid selected from glycols, glycol ethers and mixtures is present in the aqueous injectable composition in an amount of 20 g to 300 g per litre.

13. The aqueous injectable nutritional supplement composition of claim 1, comprising: 1 g to 5 g of cyanocobalamin, hydroxocobalamin or mixture thereof per litre; 10 to 60 g of zinc per litre; 10 g to 30 g of copper per litre; and 50 g to 200 g water-soluble liquid selected from glycols, glycol ethers and mixtures thereof per litre of aqueous injectable composition.

14. The aqueous injectable nutritional supplement composition of claim 1, wherein the EDTA complex is formed with di-sodium EDTA, di-potassium EDTA, di-alkyl ammonium EDTA, di-ammonium EDTA or mixture thereof.

15. The aqueous injectable nutritional supplement composition of claim 1, comprising 1 g to 10 g selenium per litre of aqueous injectable composition.

16. The aqueous injectable nutritional supplement composition of claim 1, comprising: 2 g to 4 g cyanocobalamin or hydroxocobalamin acetate per litre 20 g to 45 g of zinc per litre; 10 g to 20 g copper per litre; 3 g to 10 g selenium per litre; and 80 to 180 g propylene glycol per litre.

17. The aqueous injectable nutritional supplement composition of claim 1, wherein the pH of the aqueous injectable composition is in the range 5 to 8.

18. The aqueous injectable nutritional supplement composition of claim 1 comprising a water content of 40% to 80% v/v.

19. A method of supplementing the nutritional status of livestock comprising administering by subcutaneous injection to the livestock an aqueous injectable nutritional supplement composition according to claim 1.

20. The method of claim 19 wherein the livestock are ruminant animals.

21. The method of claim 19, wherein the livestock are administered a dose of the aqueous injectable composition of between 2 ml to 20 ml.

22. The aqueous injectable nutritional supplement of claim 1, comprising 20 g to 100 g per litre of zinc, copper or mixture thereof.

Description

DETAILED DESCRIPTION

[0023] We have found that the presence of a water-soluble liquid selected from glycols, glycol ethers and mixtures thereof has a very significant effect in stabilising vitamin B.sub.12 against degradation in an aqueous solution in the presence of the EDTA complexes of trace elements. The presence of significant amounts of trace element complexes was found to exacerbate degradation of vitamin B.sub.12 which could not be effectively controlled by methods normally used in formulation injectable vitamin B.sub.12 compositions.

[0024] Strategies previously adopted for stabilising vitamin B.sub.12 against degradation such as providing an acidic pH, the addition of aluminium sulfate or cysteine hydrochloride and formation of micelles were found to be ineffective in the presence of the high concentration of EDTA complexes of the trace elements.

[0025] The present inventor found that glycols and glycol ethers and in particular propylene glycol provided an unexpected stability of the vitamin B.sub.12 in the presence of the even high concentration of trace element complexes.

[0026] The term “glycol” refers to any of a group of alcohols containing two hydroxyl groups, particularly including propylene glycol (PG) (propane 1,2-diol) and propane-1,3-diol.

[0027] The term “glycol ether” refers to a compound in which one or both of the hydroxyl groups of a diol or condensate thereof are etherified. The term includes both monoglycol ethers and polyglycol ethers. Typically glycol ethers are of formula HO—[A—O].sub.n—R, wherein A represents ethylene or propylene moieties, R represents alkyl moieties of 1 to 4 carbon atoms, and n can assume values between 1 and 8, preferably between 1 and 4. Examples of glycol ethers include ethylene glycol monoisopropyl ether, ethylene glycol monobutyl ether, ethylene glycol monoisobutyl ether, ethylene glycol monoallyl ether, diethylene glycol monomethyl ether, diethylene glycol monoisopropyl ether, diethylene glycol monobutyl ether, diethylene glycol monoisobutyl ether, diethylene glycol monobenzyl ether, diethylene glycol dimethyl ether, diethylene glycol diethyl ether, diethylene glycol methyl ethyl ether, triethylene glycol monomethyl ether, triethylene glycol monobutyl ether, triethylene glycol dimethyl ether, polyethylene glycol monomethyl ether, propylene glycol monomethyl ether, propylene glycol monopropyl ether, dipropylene glycol monomethyl ether, tripropylene glycol monomethyl ether and 3-methoxy-3-methyl-1-butanol.

[0028] Preferred glycols and glycol ethers are water-soluble liquids, typically having molecular weight no more than 400, preferably no more than 300, particularly no more than 200 such as no more than 150. The more preferred water-soluble liquid selected from glycols and glycol ethers are propylene glycol and polyethylene glycol of molecular weight no more than 300.

[0029] Throughout the description and the claims of this specification the word “comprise” and variations of the word, such as “comprising” and “comprises” is not intended to exclude other additives, components, integers or steps.

[0030] The aqueous injectable nutritional supplement comprises an EDTA complex of one or more trace elements. Generally, the one or more trace elements will comprise at least one of zinc, copper, manganese, selenium and chromium. Preferably the trace elements comprise Zinc, copper or mixture thereof. In one embodiment the trace elements comprise zinc, copper and selenium. The amount of the trace element referred to herein is based on the element itself and not the whole trace element complex or mixture of complexes.

[0031] The aqueous injectable nutritional supplement composition will typically comprise the trace elements in an amount of 1 g to 100 g per litre of aqueous injectable composition. Preferred amounts of trace elements are in the range of from 20 g to 100 g such as 30 g/L to 100 g/L, 40 g/L to 100 g/L, 50 g/L to 100 g/L or 60 g/L to 100 g/L. Accordingly in one embodiment the trace elements comprise 1 g to 100 g per litre of zinc, copper or mixture thereof preferably 20 g to 100 g such as 30 g/L to 100 g/L, 40 g/L to 100 g/L, 50 g/L to 100 g/L or 60 g/L to 100 g/L. The total trace element content is typically up to 100 g/L of composition.

[0032] In one embodiment the content of 10 g to 60 g of zinc per litre, 5 g to 30 g of copper per litre such as 10 g to 30 g copper per litre or 10 g to 60 g of zinc per litre and 5 g to 30 g of copper per litre such as 10 g to 30 g copper per litre

[0033] The vitamin B.sub.12 may be in a range of known forms. For example, the vitamin B.sub.12 may comprise at least one of cyanocobalamin, hydroxocobalamin, hydroxocobalamin acetate, methylcobalamin and aquacobalamin and mixtures thereof. The amount of the vitamin B12 is typically 0.5 g to 8 g such as 1 g to 8 g per litre of aqueous injectable composition or 1 g to 5 g per litre of aqueous injectable composition.

[0034] The water-soluble liquid selected from glycols, glycol ethers and mixtures thereof is typically present in the aqueous injectable composition in an amount of 20 g to 300 g per litre preferably 50 g to 250 g per litre, more preferably 80 to 250 g per litre such as 100 g/L to 250 g/L or 120 to 200 g/L of the aqueous injectable composition.

[0035] In one set of embodiments the aqueous injectable supplement composition comprises: [0036] 1 g to 5 g of cyanocobalamin, hydroxocobalamin or mixture thereof per litre; [0037] 10 g to 60 g of zinc per litre; [0038] 5 g to 30 g of copper per litre such as 10 g to 30 g; [0039] 1 g to 10 g selenium per litre; and [0040] 50 g to 200 g per litre of water-soluble liquid selected from glycols, glycol ethers and mixtures thereof, preferably 50 g to 200 g propylene glycol per litre.

[0041] The aqueous injectable nutritional supplement composition comprises an EDTA complex of one or more trace elements. The one or more EDTA complexes may be formed with any of a range of EDTA forms such as di-sodium EDTA, di-potassium EDTA, di-alkyl ammonium EDTA, di-ammonium EDTA or combination of two or more thereof. Typically, we have found it convenient to use the di-sodium EDTA in forming the trace element complexes.

[0042] Selenium deficiency in many pastures in countries such as Australia can severely inhibit growth of livestock and even result in death. The aqueous injectable nutritional supplement composition preferably comprises 1 g to 10 g selenium per litre of aqueous injectable composition such as 2 g to 8 g per litre.

[0043] In a further embodiment the aqueous injectable nutritional supplement composition comprises: [0044] 2 g to 4 g cobalamin or hydroxocobalamin acetate per litre [0045] 20 g to 45 g of zinc per litre; [0046] 10 g to 20 g copper per litre; [0047] 2 g to 10 g selenium per litre; and [0048] 80 g to 180 g per litre of water-soluble liquid selected from glycols, glycol ethers and mixtures thereof, preferably 50 g to 200 g propylene glycol per litre.

[0049] The pH of the aqueous injectable nutritional supplement composition is not narrowly critical and does not have a significant effect on the degradation of the vitamin B12 in the presence of the trace element complexes. The pH is preferably the range 5 to 8 such as 6 to 8, 6.5 to 7.5 or about 7.

[0050] In embodiments of the invention in which the pH control is to provide a tight range the composition may be buffered using a suitable buffer such as a citrate, phosphate, trimethamine (also called tris buffer) or acetate buffer. Phosphate buffers, such as sodium phosphate, potassium phosphate and mixtures thereof, are preferred.

[0051] The water content of the aqueous injectable nutritional supplement composition in one set of embodiments is 40% to 80% v/v

[0052] In a further embodiment the invention provides a method of supplementing the nutritional status of livestock comprising administering by subcutaneous injection to the livestock the aqueous injectable nutritional supplement composition as hereinbefore described.

[0053] The method is particularly useful in providing nutritional supplementation to ruminant animals especially bovine animals.

[0054] The invention will now be described with reference to the following examples. It is to be understood that the examples are provided by way of illustration of the invention and that they are in no way limiting to the scope of the invention.

EXAMPLES

Comparative Example 1

[0055] A formulation, the same as a purportedly stable, commercially available and registered solution of vitamin B.sub.12 plus selenium was prepared.

[0056] The formulation contained the components in the amounts shown in Table 1.

TABLE-US-00001 TABLE 1 Elemental Compound g/L g/L Chlorocresol 1 Sodium Acetate 1 Sodium Chloride 2.67 Hydroxocobalamin Acetate (Vit B.sub.12) 1.8 Cyanocobalamin (Vit B.sub.12) 0.2 Acetic Acid 1.09 Sodium Selenate 5.0 12 Water to volume

[0057] To the formulation of Table 1, water-soluble disodium Copper EDTA and disodium Zinc EDTA, were successfully added at the rate of 107 g/L and 285 g/L respectively and then topped up with water to the required volume.

[0058] This provided a final formulation shown in Table 2.

TABLE-US-00002 TABLE 2 Elemental Compound g/L g/L Chlorocresol 1 Sodium Acetate 1 Sodium Chloride 2.67 Hydroxocobalamin Acetate (Vit B.sub.12) 1.8 Cyanocobalamin (Vit B.sub.12) 0.2 Acetic Acid 1.09 Sodium Selenate 5.0 12 Disodium Copper EDTA 15 107 Disodium Zinc EDTA 40 285 Selenium Selenate 5 8 Water to volume

[0059] The formulation was fully dissolved with no evidence of precipitate.

[0060] This formulation was sent to Chemical Analysis, and analysed at 0 time and placed on stability storage at 5° C. and 40° C. (75% RH).

[0061] The results were as shown in Table 3.

TABLE-US-00003 TABLE 3 T = 0 (mg/mL) T = 1 month (mg/mL) Vitamin B.sub.12 5° C. 40° C. 5° C. 40° C. Cyanocobalamin 0.24 0.24 0.23 0.14 Hydroxocobalamin 0.49 0.49 0.26 0.03 Acetate

[0062] Due to the obvious lack of stability of the formulation the trial was abandoned.

Comparative Example 2

[0063] Formulations shown in Table 4 were prepared with single or combinations of chelated minerals (disodium EDTA) were formulated with Cyanocobalamin (CNCBL) and subjected to screening stability studies.

TABLE-US-00004 TABLE 4 Form- Components 40° C. 40° C. 40° C. ulation (g/L) pH Month 1 Month 2 Month 3 1 CNCBL 4.0 6.94 CNCBL 3.3  CNCBL 3.3 Zn 40 Water to Vol 2 CNCBL 4.0 4.4  CNCBL 3.3  CNCBL 3.3 Cu 16 Water to Vol 3 CNCBL 4.0 6.18 CNCBL 3.1  CNCBL 3.2 Zn 20.0 Cu 7.5 Water to Vol 4 CNCBL 4.0 4.32 CNCBL 3.3  CNCBL 1.8 Zn 40 Water to Vol 5 CNCBL 2.5 5.9  CNCBL 1.24 Zn 40.0 Cu 15.5 Water to Vol 6 CNCBL 2.5 5.9  CNCBL 1.47 Zn 40.0 Cu 15.5 Se 5.0 Water to Vol CNBL—Cyanocobalamin

[0064] The results shown in Table 4 demonstrate that in the presence of 40 g/L of Zinc (as disodium zinc EDTA), 15 g/L of Copper (as disodium copper EDTA) and 5.0 g/L of selenium (as selenium selenate) (in formulations 4, 5 and 6), which are the levels necessary to provide enough trace mineral in a single injection to livestock, sufficient levels of cyanocobalamin could not be maintained over time. Contrary to reported findings in the literature lowering the pH did not enhance the stability of the cyanocobalamin.

Comparative Example 3

[0065] It has been claimed that the addition of Aluminium sulphate may enhance the stability of vitamin B.sub.12 in aqueous solutions.

[0066] The solutions of Table 5 were formulated and subjected to accelerated stability testing at 40° C.

TABLE-US-00005 TABLE 5 For- Components 40° C. 40° C. 40° C. 40° C. mulation (g/L) pH Month 0 Month 1 Month 2 Month 3  7 CNCBL 2.5 5.8 CNCBL CNCBL Zn 40.0 2.0 1.06 Cu 15.5 Se 5.0 Al. Sulphate 62.5 10 CNCBL 2.5 5.1 CNCBL CNCBL CNCBL Zn 40.0 2.3 1.70 1.36 Cu 15.5 Se 5.0 Al. Sulphate 10.0

[0067] It was concluded that the addition of Aluminium sulphate did not enhance the stability of Cyanocobalamin in solutions containing high levels of trace minerals

Comparative Example 4

[0068] Micelle formulations have been used to enhance the stability of molecules that are soluble in organic solvents but break down in aqueous solutions. Solutions including organic solvents singularly or in combination and a suitable surfactant were formulated and tested for stability of the cyanocobalamin in accordance with Table 6.

TABLE-US-00006 TABLE 6 For- Components 40° C. 40° C. 40° C. 40° C. mulation (g/L) pH Month 0 Month 1 Month 2 Month 3 8 CNCBL 2.5 6.0 CNCBL CNCBL CNCBL Zn 40.0 2.31 1.31 1.09 Cu 15.5 Se 5.0 G Formal* 100.0 ECT 80** 10.0

[0069] GFormal*—Glycerol Formal; ECT80**—Polyoxyethylene sorbitan monooleate (Tween 80); PG—propylene glycol.

[0070] It was concluded that these attempts to stabilise the vitamin B.sub.12 in a micelle formulation were not successful.

Example 1

[0071] In the process of combining organic solvents into the aqueous formulations of vitamin B.sub.12 and high concentrations of trace minerals (during attempts to produce micelles containing the vitamin B.sub.12) it was surprisingly discovered that by adding propylene glycol alone increased the stability of vitamin B.sub.12 in a solution containing high amounts of trace minerals

[0072] The formulations and stability results are shown in Table 7.

TABLE-US-00007 TABLE 7 Components 40° C. 40° C. 40° C. 40° C. 40° C. 40° C. Formulation (g/L) Month 0 Month 1 Month 2 Month 3 Month 4 Month 5 11 CNCBL 3.0 CNCBL CNCBL CNCBL (Low Zn 30.0 3.12 2.36 2.36 mineral Cu 12 content) Se 5.0 PG 100 T80 2.0 12 CNCBL 3.0 CNCBL CNCBL CNCBL Zn 40.0 2.42 2.38 2.33 Cu 15 Se 5.0 PG 100 T80 2.0 26 CNCBL 2.50 CNCBL CNCBL CNCBL CNCBL (23.4.18) Zn 40.0 2.45 2.4 2.35 2.40 Cu 20.0 Se 5.0 PG 150 26 CNCBL 2.50 CNCBL CNCBL CNCBL CNCBL CNCBL (23.6.18) Zn 40.0 2.58 2.3 2.22 2.15 2.14 Cu 20.0 Se 5.0 PG 150

[0073] The following Trial formulations were initially studied.

[0074] T80—Polyoxyethylene sorbitan monooleate (Tween 80)

Example 2

[0075] A further study mimicking commercial production and packaging was implemented and is reported below:

[0076] Method of Manufacture:

[0077] Part A [0078] 1. Load warm water (40° C.) to mixing tank. [0079] 2. Load disodium EDTA dihydrate powder to water and stir. [0080] 3. Load Zinc Oxide powder to suspension and stir. [0081] 4. Load Copper Hydroxide to suspension and stir. [0082] 5. Continue stirring until all solids have dissolved; about 2 hours will be required. [0083] 6. The reaction can be accelerated by heating the suspension at 40° C.; a clear blue solution will be obtained. [0084] 7. The pH of the solution at this stage should be 6.5-7.0. [0085] 8. Load Sodium Selenate to batch and stir till dissolved.

[0086] Part B [0087] 9. Load Propylene Glycol to small reactor. [0088] 10. Load Cyanocobalamin powder to Propylene Glycol and stir till dissolved; slight warming to 40 deg. C will assist dissolution [0089] 11. Load all of Part B to Part A in main tank and stir the solution till uniform. [0090] 12. Measure the visible absorbance of the solution; 2% solution should have Absorbance at 550 nm>0.31 [0091] 13. Filter the solution through 10 micron gauze to remove any insoluble solids, when transferring to holding tank. [0092] 14. Filter product through 0.25 micron filter into sterile HDPE/LDPE “flow-pack” containers under aseptic conditions.

[0093] Once manufactured the formulation was placed in stability testing at 40° C. The formulation is shown in Table 8 and the stability testing results are shown in Table 9 below.

TABLE-US-00008 TABLE 8 Components Formulation (g/L) 26 (21.9.18) CNCBL 2.50 Zn 40.0 Cu 20.0 Se 5.0 PG 150

TABLE-US-00009 TABLE 9 40° C. 40° C. 40° C. 40° C. 40° C. 40° C. 40° C. 40° C. Month Month Month Month Month Month Month Month 0 1 2 3 4 5 6 7 CNCBL CNCBL CNCBL CNCBL CNCBL CNCBL CNCBL CNCBL 2.19 2.12 2.32 2.31 2.28 2.32 2.31 2.25

Example 3

Composition Containing Dipropylene Glycol Monomethyl Ether

[0094] A composition in accordance with the invention was prepared by combining premixed compositions identified as Part A and Part B below:

TABLE-US-00010 Part A Water  65.0 grams Na2H2EDTA  32.5 grams ZnO  5.0 grams Cu(OH)2  2.6 grams Na Selenate  1.2 grams Total 106.3 grams

TABLE-US-00011 Part B “DOWINOL” DPM 14.0 grams Propylene Glycol  3.0 grams CN-Cbl 0.28 grams Total 17.3 grams

[0095] Note: DOWINOL DPM is dipropylene glycol monomethyl ether (DOWINOL is a trademark of the Dow Chemical Company).

[0096] Part B was added to Part A to provide a total combined batch weight of 123.6 grams.

[0097] The composition was subject to stability testing at 40° C. for 100 days and the analytical results are shown in Table 10 below.

TABLE-US-00012 TABLE 10 0 Days 30 Days 80 Days 100 Days SG (g/L) 1200 1200 1200 pH 6.16 6.36 6.47 Cu (g/L) 14.8 15.3 15.4 15.6 CN-Cbl (g/L 2.33 2.41 2.4 2.45 Zn (g/L) 40 (calculated) Se (g/L 5 (calculated)

[0098] The results show no loss of Vitamin B12 after 100 days at 40° C.

Example 4

Composition Containing Triethylene Glycol (PEG 150)

[0099] A composition in accordance with the invention was prepared by combining premixed compositions identified as Part A and Part B below:

TABLE-US-00013 Part A Water 52.26 g Na.sub.2H.sub.2EDTA  27.4 g ZnO  5.1 g Cu(OH).sub.2  1.0 g Total 87.06 g

TABLE-US-00014 Part B Triethylene Glycol 12.7 g Cn-Cbl  0.2 g

[0100] Part B was added to Part A to provide a total combined batch weight of 100 g.

[0101] The composition was subject to stability testing at 40° C. and the analytical results are as shown in Table 11.

TABLE-US-00015 TABLE 11 0 Days 30 Days 60 Days SG (g/L) 1235 pH 6.72 6.46 Cu (g/L) 17.2 17.4 17.4 CN-Cbl (g/L 2.61 2.56 2.55 Zn (g/L) 40 (calculated) Se (g/L 5 (calculated)

[0102] The results show virtually no loss of Vitamin B12 after 60 days at 40° C.

Example 5

Composition Containing Polyethylene Glycol MW 200

[0103] A composition in accordance with the invention was prepared by combining premixed compositions identified as Part A and Part B below:

TABLE-US-00016 Part A Water 523.6 g Na.sub.2H.sub.2EDTA 274.0 g ZnO  41.0 g Cu(OH).sub.2  22.0 g Total 870.6 g

TABLE-US-00017 Part B PEG 200   127 g Cn-Cbl  2.4 g Total 129.4 g

[0104] Part B was added to Part A to provide a total combined batch weight of 1000 g.

[0105] The composition was subject to stability testing at 40° C. and the analytical results are as shown Table 12.

TABLE-US-00018 TABLE 12 0 Days 30 Days 60 Days SG (g/L) pH 6.21 Cu (g/L) 16.8 16.7 16.8 CN-Cbl (g/L 2.71 2.65 2.65 Zn (g/L) 40 (calculated) Se (g/L 5 (calculated)

[0106] The results show Vitamin B12 stable after 60 days at 40° C.

Example 6

Composition Manufactured Under Commercial Conditions in a Commercial Manufacturing Facility

[0107] Some laboratory scale formulations may differ in performance and stability to larger scale formulations manufactured under commercial conditions in a commercial manufacturing plant

[0108] A composition in accordance with the invention in a commercial scale facility was manufactured by combining premixed compositions identified as Part A and Part B below. The composition was then filtered through a 0.2μ filter and aseptically filled into 500 mL HDPE/LDPE flow-pack containers

TABLE-US-00019 Part A Water 7.7895 L Na.sub.2H.sub.2EDTA 4.1670 Kg ZnO 0.6150 Kg Cu(OH).sub.2 0.3375 Kg

TABLE-US-00020 Part B Polyethylene Glycol 200 1.7250 Benzyl Alcohol  0.18 Kg Cn-Cbl 0.0360 Kg Total  14.85 Kg

[0109] After 3 months at ambient temperature the composition was subjected to storage at 40° C.

[0110] The analytical results are as shown in Table 13.

TABLE-US-00021 TABLE 13 Ambient 40° C. Month 0 Month 3 Month 4 Month 5 Month 6 Month 7 pH 7.07 7.11 7.29 7.41 6.93 CnCbl 2.06 1.99 1.97 1.95 2.03 2.02 Copper 16.3 16.2 16.5 16.2 16.5 16.3

[0111] The results show that a composition in accordance with the invention manufactured in a commercial scale facility under commercial conditions including sterilization by filtration and packaging remained stable with respect to Vitamin B12 for 7 months at 40° C.

Example 7

Composition with the Addition of a pH Buffer

[0112] It may be advantageous to ensure that the pH of a composition in accordance with the invention was held between a specified narrow range. Accordingly, a phosphate/potassium buffer was added to the above composition between month 5 and month 6 in an amount calculated to hold the pH between pH 6.5 and pH 7.0.

[0113] The amount added was as follows:

TABLE-US-00022 Disodium phosphate 10.6 g Monopotassium phosphate 11.1 g

[0114] Analytical results were as shown in Table 14.

TABLE-US-00023 TABLE 14 Month 0 Month 3 Month 4 Month 5 Month 6 Month 7 pH 7.07 7.11 7.29 7.41 6.93 6.85

[0115] The results showed that very small amount of buffer can stabilize the pH in a desired narrow range.