Determination of Menaquinone-7 by a Simplified Reversed Phase- HPLC Method

Author(s): Dinali Ranmadugala, Megan Grainger, Merilyn Manley-Harris, Aydin Berenjian*.

Journal Name: Current Pharmaceutical Biotechnology

Volume 19 , Issue 8 , 2018

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Abstract:

Background: An efficient and accurate HPLC method was developed for the determination of menaquinone-7 (MK-7) in microbial fermentation using 2-propanol and n-hexane as extraction solvents as well as the eluent.

Methods: Extraction was carried out with 2-propanol and n-hexane (2:1, v/v) after enzymatic hydrolysis with 1% (w/v) lipase and ethanol water treatment prior to quantification in order to remove interfering lipids and denatured proteins. Chromatographic separation of MK-7 was accomplished isocratically on a C 18 Gemini column using a mobile phase mixture of 2- propanol: n-hexane (2:1, v/v) with a flow rate of 0.5 mL/min. UV detection was carried out from 200-400 nm and the chromatogram was extracted at a wavelength of 248 nm. A linear response was shown by the method with a coefficient of determination (R2) value of 0.9982.

Results: The recoveries of MK-7 were greater than 94% and the intra and inter day R.S.D values were less than 2%, demonstrating the accuracy of the method. The lower limit of detection (LOD) and the limit of quantification (LOQ) were 0.1 µg/mL and 0.29 µg/mL, respectively.

Conclusion: The general usefulness of the described method is demonstrated by the application of this method in the analysis of MK-7 from Bacillus species. Under these conditions, the analysis of MK-7 was achieved in less than 8 minutes with a retention time of 7.19 ± 0.1 minutes.

Keywords: High-performance liquid chromatography, Menaquinone-7, bacterial fermentation, interfering substances, inoculum preparation, vitamin K analysis.

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Article Details

VOLUME: 19
ISSUE: 8
Year: 2018
Page: [664 - 673]
Pages: 10
DOI: 10.2174/1389201019666180828090637
Price: $58

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