Green fast and simple UPLC-ESI-MRM/MS method for determination of trace water-soluble vitamins in honey: Greenness assessment using GAPI and analytical eco-scale

Abstract

Honey is one of the most commonly used natural products for nutritional and therapeutic purposes. Besides sugars, honey is characterized by complex chemical composition of minerals, polyphenols, water soluble vitamins, carotenoids and organic acids. In the present study, nine water soluble vitamins; C (L-ascorbic acid), B1 (thiamine), B2 (Riboflavin), B3H (nicotinic acid), B5 (D-pantothenic acid), B6 (pyridoxine), B7 (Biotin), B9 (Folic acid) and B12 (Cyanocobalamin) are determined simultaneously for the first time using an ultra-performance liquid chromatography-electrospray ionization multiple reaction monitoring/mass spectrometry (UPLC-ESI-MRM/MS) method coupled with a single simple sample preparation step. The simplicity and sensitivity of the proposed method permit the simultaneous determination of all nine water soluble vitamins including vitamins B7 and B12 that are present in trace amounts in various real honey samples. Complete chromatographic separation is achieved in 3 mins on a C18 column with a green mobile phase composed of a mixture of aqueous formic acid (0.1%, v/v) and methanol in gradient elution mode at a flow rate of 0.4 mLmin−1. The method is fully validated according to ICH guidelines. The greenness of our proposed method is assessed by comparing it to other proposed methods in terms of sample preparation, instrument energy consumption, consuming hazardous reagents and waste generation using Analytical Eco-scale and GAPI (Green Analytical Procedure Index). Results proved the superior greenness of our proposed method over the other reported ones. The proposed method is applied to twelve samples from six different monofloral honey types. Quantitation of the water-soluble vitamins content revealed concentrations ranging from 0.63 to 936.98 mgKg−1 in the analyzed honey samples with the successful determination of vitamins B7 and B12 in trace amounts. The highest vitamin content detected was for vitamin B3 in Jujube-tree honey while the lowest vitamin content was for vitamin B12 in Orange-tree honey. A marked relation was observed between the botanical origin and the vitamins’ concentrations of the studied honey samples. In addition, factors such as temperature, moisture, light and pH during storage greatly affect the vitamins content of honey. The developed method is intended to serve as a reliable, accurate, fast and green tool for the routine analysis of water-soluble vitamins in honey samples for quality control and research purposes.