Vitamin B6 (Pyridoxine)

High Performance Liquid Chromatography (HPLC) Method for Analysis of Water-Soluble Vitamins (B1, B2, B3, B6) on Primesep 100 by SIELC Technologies

Separation type: Liquid Chromatography Mixed-mode SIELC Technologies

 High Performance Liquid Chromatography (HPLC) Method for Analysis of Vitamin B1 (Thiamine), Vitamin B2 (Riboflavin), Nicotinamide, Vitamin B6 (Pyridoxine), Pyridoxamine, Pyridoxal Phosphate

Water-Soluble Vitamins (B1, B2, B3, B6) can be retained, separated and analyzed using a Primesep 100 mixed-mode stationary phase column. The analysis employs an gradient method with a simple mobile phase comprising water, acetonitrile (MeCN), and sulfuric acid as a buffer. This method allows for detection using UV 300 nm.

You can find detailed UV spectra of Vitamin B1 (Thiamine), Vitamin B2 (Riboflavin), Nicotinamide, Vitamin B6 (Pyridoxine), Pyridoxamine, Pyridoxal Phosphate and information about its various lambda maxima by visiting the following links: Vitamins (B1, B2, B3, B6)

High Performance Liquid Chromatography (HPLC) Method for Analysis of Nicotinamide, Vitamin B6 (Pyridoxine) on Primesep 100 by SIELC Technologies

Separation type: Liquid Chromatography Mixed-mode SIELC Technologies

 High Performance Liquid Chromatography (HPLC) Method for Analysis of Nicotinamide, Vitamin B6 (Pyridoxine)

Nicotinamide is a specific form of vitamin B3, an essential nutrient that is crucial for various bodily functions. Along with niacin (nicotinic acid), nicotinamide is one of the primary forms of vitamin B3 that the body uses. Both of these forms are converted into coenzymes NAD+ and NADP+, which play vital roles in energy production, DNA repair, and cellular communication. This conversion is essential for maintaining the body’s metabolic processes, highlighting the importance of nicotinamide as a form of vitamin B3.

Pyridoxine is a form of vitamin B6, which is essential for numerous biochemical processes in the body. Vitamin B6 exists in three natural forms—pyridoxine, pyridoxal, and pyridoxamine—all of which are converted into the active coenzyme pyridoxal 5′-phosphate (PLP). This active form is critical for amino acid metabolism, neurotransmitter synthesis, and hemoglobin production. As a form of vitamin B6, pyridoxine is key to maintaining proper neurological and immune function, ensuring that the body can perform a wide range of essential functions.

Nicotinamide, Vitamin B6 (Pyridoxine) can be retained, separated and analyzed using a Primesep 100 mixed-mode stationary phase column. The analysis employs an isocratic method with a simple mobile phase comprising water, acetonitrile (MeCN), and sulfuric acid as a buffer. This method allows for detection using UV 270 nm.

You can find detailed UV spectra of Nicotinamide, Vitamin B6 (Pyridoxine) and information about its various lambda maxima by visiting the following links: UV-Vis Spectrum of Nicotinamide, UV-Vis Spectrum of Pyridoxine.

For optimal results in HPLC analysis, it is recommended to measure absorbance at a wavelength that matches the absorption maximum of the compound(s) being analyzed. The UV spectrum shown can assist in selecting an appropriate wavelength for your analysis. Please note that certain mobile phases and buffers may block wavelengths below 230 nm, rendering absorbance measurement at these wavelengths ineffective. If detection below 230 nm is required, it is recommended to use acetonitrile and water as low UV-transparent mobile phases, with phosphoric acid and its salts, sulfuric acid, and TFA as buffers.
For some compounds, the UV-Vis Spectrum is affected by the pH of the mobile phase. The spectra presented here are measured with an acidic mobile phase that has a pH of 3 or lower.

Separation type: Liquid Chromatography Mixed-mode

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High Performance Liquid Chromatography (HPLC) Method for Analysis of Pyridoxine Hydrochloride (Vitamin B6) and Benfotiamine

Pyridoxine Hydrochloride (also known as Vitamin B6), is a naturally occurring vitamin that can be found in a wide variety of common foods, such as meat, grains, and avocados. Benfotiamine is a synthetic form of Vitamin B1 that has shown early signs of treating nerve damage, Alzheimer’s, alcohol dependence. They can both be found in Milgamma 100 tablets, a medicine prescribed to treat Vitamin B1 and B6 deficiency.
These two vitamins can be detected in the low UV regime. Using a Primesep S normal-phase column and a mobile phase consisting of water and acetonitrile (MeCN) with a phosphoric acid (H3PO4) buffer, Pyridoxine Hydrochloride and Benfotiamine can be retained, separated, and analyzed. This analysis method can be UV detected at 280 nm.

Pyridoxine, also known as Vitamin B6, is an essential nutrient required by the body to produce red blood cells and for proper nerve functioning. Doxylamine is a common treatment for insomnia. When taken together, they can treat morning sickness in pregnant women.

Pyridoxine and Doxylamine can be retained, separated, and detected on a normal-phase Primsep S using an isocratic analytical method with a simple mobile phase of water, Acetonitrile, and an Ammonium Formate buffer. This analysis method can be detected in the low UV regime at 270 nm.

Application Notes: Pyridoxine is part of the vitamin B complex group. Pyridoxine is important in the body’s daily function as it regulates many enzymatic reactions. The USP HPLC method for the separation of pyridoxine was developed on Legacy L1 column according to the US Pharmacopeia methodology. L1 classification is assigned to reversed-phase HPLC column containing C18 ligand. Support for the material is spherical silica gel with particles size 3-10 um and pore size of 100-120A. Resolution between critical pairs corresponds to rules and specifications of UPS.  

Application Columns: Legacy L1 C18 HPLC column

Application compounds: Pyridoxine

Mobile phase: AcOH/MeOH/H2O/Hexanesulfonate (2/47/153/1.2)

Detection technique: UV

Reference: USP35- NF30

HPLC method for separation of active ingredients of drug/supplemental composition was developed on an Obelisc R trimodal HPLC column. Compounds are retained by combination of reversed-phase, cation-exchange and anion-exchange mechanisms. Compounds are well separated, and method can be used for quantitation of pyridoxine, ascorbic acid, niacinamide, pantothenic acid, caffeine and riboflavin in a mixture or as separate compounds in various complex mixtures. Various detection techniques can be applied for quantitation (ELSD, UV, LC/MS, Corona). This HPLC method can be adopted as general approach for analysis of active drug components in various formulations.

Vitamin C (ascorbic acid) and Vitamins Group B are separated on Obelisc N mixed-mode column. Method can be used in quantitation and determination of polar vitamins in various formulations and dietary supplements. HPLC method can be based on UV, Evaporative Light Scattering Detection (ESLD), RI or MS detection. Effect of sample matrix can be eliminated by changing mobile phase conditions. Buffer concentration, buffer pH and amount of ACN will affect every vitamin differently due to difference in polar and ionic properties.

Separation of vitamins group B is achieved on Obelisc N column in HILIC mixed-mode. Vitamins of this group are different in polarity and ionic properties. Retention and separation is achieved by optimization of amount of ACN, buffer and buffer pH. Combination of UV and ELSD detection is used to monitor HPLC separation. B vitamins are water-soluble vitamins that play an important role in cell metabolism. Supplements containing all six are generally referred to as a vitamin B complex. Individual B vitamin supplements are referred to by the specific name of each vitamin. This method can be used to analyze individual B vitamins as well as vitamin B complex. Isolation of impurities as well as degradation products is possible by preparative chromatography.

Primesep C separates B vitamins with baseline resolution by a combination of cation exchange, complex formation, and hydrophobic interactions. Vitamin B1 (thiamine), vitamin B2 (riboflavin), and vitamin B6 (pyridoxine) are separated with a mobile phase of water, acetonitrile (MeCN, ACN) and triethylamine (TEA) phosphate with UV detection at 280 nm.