Effect of Various Halide Salts on the Incompatibility of Cyanocobalamin and Ascorbic Acid in Aqueous Solution

Abstract

Cyanocobalamin (VB12) and ascorbic acid (VC), both water-soluble vitamins, are essential nutrients involved in many physiological functions. These vitamins are extensively used in many pharmaceutical preparations and dietary supplements. An incompatibility of VB12 and VC in aqueous solutions has been reported. [1][2][3][4][5][6][7] However, it has not been fully elucidated how to stabilize these vitamins in a mixed solution yet. Many researchers have investigated the effect of VC on the stability of VB12 or cobalamin analogues in solution. [8][9][10][11][12] Most of them have focused only on the VB12 degradation, but not on VC. While Bartilucci et al. 6,7) have reported the interaction of VB12 and VC mixed condition in aqueous solutions evaluated by less sensitive methods, microbiological method for VB12 and titration for VC.In the present study, we determined the stabilities of VB12 and VC in acetate buffer using high performance liquid chromatography (HPLC), which shows better accuracy and precision than microbiological method and titration. The effects of halide salts on the stabilities of these vitamins in the buffer were also studied for the improvement of their stability. ExperimentalMaterials and Reagents VC and VB12 regulated by Japanese Pharmacopoeia were obtained from BASF Takeda Vitamins (Tokyo, Japan) and Aventis Pharma S.A. (Alsace, France), respectively. All other chemicals were purchased commercially. Water purified through a Milli-Q Labo (Millipore, Bedford, MA, U.S.A.) water system was used for all dilutions and sample preparations.Determination of VB12 Using HPLC An LC-10AVP system (Shimadzu, Kyoto, Japan) equipped with SIL-10ADVP auto-injector, SPD-10AVVP detector, LC-10ADVP pump, DGU-14A degasser, CTO-10ACVP column oven and C-R6A data processor was used to determine VB12 in the solutions. The HPLC conditions were as follows: column, 150ϫ4.6 mm TSK gel ODS-80TsQA (Tosoh, Tokyo, Japan); column temperature, 25°C; detection wavelength, 361 nm; mobile phase, acetonitrile-50 mM acetate buffer, pH 4.5 (13 : 87, v/v); flow rate, 1.0 ml/min; injection volume, 50 ml. Sample mixtures were directly injected into HPLC.Determination of VC Using HPLC An LC-10AVP system, the same composition system as described above, was used. The HPLC conditions were as follows: column, 150ϫ6 mm YMC pack ODS-AQ312 (YMC, Kyoto, Japan); column temperature, 25°C; detection wavelength, 290 nm; mobile phase, water-methanol-200 mM phosphate solution, pH 3.5 (8 : 1 : 1, v/v/v); flow rate, 1.0 ml/min; injection volume, 5 ml. Samples for HPLC analysis were prepared as follows: 0.3 ml of sample mixtures were transferred to vials for HPLC followed by adding 0.3 ml of 3% metaphosphoric acid containing 2% hydrochloric acid. The mixtures were analyzed immediately by HPLC.Influence of VC on VB12 Stability Various concentrations of VC (0, 1, or 5 mM) with 0.25 mM VB12 in 100 mM ammonium acetate-acetic acid buffer (pH 4.8) were prepared in a 12-ml brown glass vial with cap, and the mixtures were kept at room temperature for 8 h. The conce...

Effect of Ascorbic Acid on the Degradation of Cyanocobalamin and Hydroxocobalamin in Aqueous Solution: A Kinetic Study

“…Similar reactions occur on the chemical degradation of B 12 and B 12b in the presence of ascorbic acid (AH 2 ) (12)(13)(14) and may be expressed as follows: The effect of thiamine (15,16), nicotinamide (17,18), and riboflavin (19)(20)(21)(22) on the photolysis of B 12 and B 12b has been investigated. Several studies have been conducted on the incompatibility and interaction of B 12 and AH 2 that lead to the loss of B 12 in aqueous solution (12,13,(23)(24)(25)(26)(27). B 12b has been found to be less stable than B 12 in the presence of AH 2 (28)(29)(30) and is destroyed to form unknown oxidation products ( 9 , 12,31, 32).…”

'Bone Broth is So Last Year - I Vape my Vitamins now' - The vaporisation of bioactive molecules in e-cigarette devices.

“…Also contributing to the degradation of bioactives, vitamins in particular, is exposure to air, UV light or temperature. 182,187,188,199 It is hard to see how these vaporisers could be modified to limit or prevent bioactive degradation, such as decrease of temperature by elimination the heating element of the device as it is required to produce the vapour, while many of these vitamins (thiamine for example) begin to degrade at temperatures as low as 50 °C. 172,200 Future degradation studies should examine the effect caused by the combined presence of other bioactive ingredients and monitor the speed of degradation and attempt to characterise some of the degradation products generated.…”

Vitamin B12 in Foods, Food Supplements, and Medicines—A Review of Its Role and Properties with a Focus on Its Stability

“…Ichikawa et al showed that the incompatibility of vitamin B 12 and ascorbic acid can be also overcome by the addition of various halide salts. They demonstrated that the presence of potassium, magnesium, and calcium halides improves the stability of both CNCbl and ascorbic acid in the pH range of 3.5-5.3 and that their stabilization effect increases with an increase in the concentrations and the atomic number of the halide anion Cl − < Br − < I − ) [242].…”