Determination of Vitamin B6 Content in Vitamin B6 Tablets

As suggested by the name, the primary ingredient of Vitamin B6 tablets is Vitamin B6. These tablets are suited for the treatment of Vitamin B6 deficiencies and isoniazid poisoning, but can also be used to treat the vomiting and seborrheic dermatitis caused by anticancer drugs and radiation sickness during pregnancy. They can be used to treat infant seizures, or be given to pregnant women to prevent infant seizures.

As the primary ingredient in such tablets, and thus the primary factor in the dosage size. As such, accurate determination of the Vitamin B6 content is of vital important to those performing research, or to those involved at the product quality assurance level. 

This report describes the method by which of the L-3000 HPLC System can be used to accurately and reliably determine the Vitamin B6 content in Vitamin B6 tablets.

Vitamin B6 Content Assay

The following equipment were used in the determination of IgG content in bovine colostrum experiment: A 5-piece Dionamix HPLC system (Consisting of an L-3120 Solvent Organizer, an L-3220 Binary High-Pressure Gradient Pump, an L-3400 Column Thermostat, an L-3300 Autosampler, and an L-3500 UV/Visible Detector), a 5µm, 4.6mmx250mm Dionamix Compass C18, a Millipore Milli-Q ultrapure water purification system, a Sartorius Germany BT 124S scale, a set of Eppendorf 10-100µL and 10-1000µL pipettes, and a Tianjin Autoscience Ultrasound. 

The experiment ran for 15 minutes, and was carried out under the following parameters: A flow rate of 1.0mL/min, an injection volume of 3µL, a detection wavelength of 291nm, and a column temperature of 40°C.

Mobile Phase

The mobile phase was prepared from an A and B component. Component A was composed of 0.04% sodium heptane sulfonate, adjusted to a pH of 3.0 with glacial acetic acid. Component B was Methanol. The two were mixed with a 75:25 A:B ratio.

Sample Preparation

Standard Curve Solutions

0.0105g of vitamin B6 reference substance was added to a 10mL volumetric flask, and mobile phase was added to the mark to produce a solution with a concentration of 1.06mg/mL (Stock Solution). The process was repeated with 2mL and 1mL of the Stock Solution to produce solutions with concentrations of 212.00µg/mL (Standard Solution (1)) and 106µg/mL (2). The process was repeated with 1ml of (1) to produce a solution with a concentration of 21.20µg/mL (3). The process was repeated with 1mL and 0.5mL of (4) to produce solutions with concentrations of 10.60µg/mL (5) and 5.30µg/mL (6)

Sample Solutions

30 Vitamin B6 tablets (1.9104g) were weighed and ground down. An amount approximately equal to 0.1g of vitamin B6 (0.6364g) was added to a 100mL volumetric flask. An appropriate amount of mobile phase was added and sonicated, after which it was diluted to the mark with mobile phase. The mix was then filtered, after which 5mL of the filtrate was added to a 50mL measuring flask, and diluted to the mark with mobile phase (Sample 1). The process was repeated, though with 0.0501g of the vitamin B6 reference substance also added to the powder before the first amount of mobile phase was added (Sample 2)

Conclusion

The L-3000 HPLC is capable of quickly and efficiently carrying out the Vitamin B6 content experiment to a high degree of accuracy, and with good qualitative and quantitative repeatability. 

This makes the Dionamix L-3000 HPLC system and method well suited to the needs of research scientists, quality control managers, industry regulators, and other similar roles that require a precise understanding of the content found within dietary supplement tablets or similar.

Applicable to research and quality assurance processes, the HPLC system and method also delivers more accurate results than other methods, and is additionally quite versatile.