Assay of Vitamin B6 (Pyridoxine Hydrochloride) Utilizing Isocratic Reversed Phase High Performance Liquid Chromatography | Chapter 02 | Advances in Applied Science and Technology Vol. 3
Aims:
To demonstrate an analysis for vitamin
B6 from commercial aqueous nutritional
drinks and solid tablets, utilizing
isocratic conditions with high performance liquid chromatography (HPLC) and UV
detection at 290 nm.
Study
Design: Vitamin B6 in the form of pyridoxine
hydrochloride is assayed by HPLC from various samples.
Place and
Duration of Study: Department of
Chemistry, Durham Science
Center, University of Nebraska, Omaha Nebraska from May to
August 2016.
Methodology:
Utilizing a reversed-phase C-18 column with eluent solvent (19% ethanol, 77%
water, 4% acetic acid), the samples were prepared in sample solvent (19%
ethanol, 81% distilled water). Detection
of vitamin B6 was
accomplished at 290
nm. Analysis of
samples was done
following solubilizing in aqueous conditions having ethanol at 10% to
20% (v/v). Column pressure at 1900 psig, rise time 0.1 with flow rate 1.0
mL/minute. Elution peak for vitamin B6 occurred con sistently at 1.6 minutes.
Nutritional samples, aqueous samples, and solid pills were prepared in aqueous
solvent with various levels of ethanol.
Results:
Levels of vitamin B6
detected were as low
as 4.4029x10-5 molar
to 7.8081x10-4 molar. Sensitivity for vitamin B6 was highest at 290
nm. Reverse phase isocratic conditions is shown to be effective for
determination of vitamin B6 in
aqueous based samples. Standard curves
applied are highly linear
in range from
zero to 7.8081x10-4 molar (y
= 112,521,145.5x + 2,818.6),
having coefficient of determination (R²= 0.9948) with very strong
positive correlation coefficient (r= 0.9974). Percent recovery of vitamin ranged from
95% to 105%. Amounts of vitamin present
in drinks from same manufacturer were consistent.
Conclusion:
Utilizing reversed phase column, isocratic solvent conditions with ethanol in
water, and a UV detector set at 290 nm is effective for determination of
vitamin B6. Ethanol - water solvent
system is effective. Vitamin B6 was found in various amou nts in nutritional
drinks tested. Percent recovery of vitamin
averaged 101% with a standard deviation of 2.4%. Pyridoxine hydrochloride was
effectively assayed from aqueous
samples, vitamin preparations,
and vitamin tablets. The
methodology presented in th is study will be useful for quality control
analysis for commercial production. Analysis methods for vitamin assay are a necessary objective to ensure
quality control of commercial products and medicinal applications.
Author(s) Details
Dr. Ronald Bartzatt
University of Nebraska,
Durham Science Center, 6001 Dodge Street, Omaha, Nebraska 68182, USA.
View Volume: https://doi.org/10.9734/bpi/aast/v3
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