Intra-Laboratory Validation of Alpha-Galactosidase Activity Measurement in Dietary Supplements.

Elena Fabris,Alessandro Nencioni,Michela Bulfoni,Emanuele Nencioni

doi:10.3390/molecules26061566

Abstract

Introduction: Alpha-galactosidase (α-Gal) is an enzyme responsible for the hydrolyzation of glycolipids and glycoprotein commonly found in dietary sources. More than 20% of the general population suffers from abdominal pain or discomfort caused by intestinal gas and by indigested or partially digested food residuals. Therefore, α-Gal is used in dietary supplements to reduce intestinal gases and help complex food digestion. Marketed enzyme-containing dietary supplements must be produced in accordance with the Food and Drug Administration (FDA) regulations for Current Good Manufacturing Practice (cGMPs). Aim: in this work we illustrated the process used to develop and validate a spectrophotometric enzymatic assay for α-Gal activity quantification in dietary supplements. Methods: The validation workflow included an initial statistical-phase optimization of materials, reagents, and conditions, and subsequently a comparative study with another fluorimetric assay. A final validation of method performance in terms of specificity, linearity, accuracy, intermediate-precision repeatability, and system precision was then executed. Results and conclusions: The proven method achieved good performance in the quantitative determination of α-Gal activity in commercial food supplements in accordance with the International Council for Harmonisation of Technical Requirements for Pharmaceuticals (ICH) guidelines and is suitable as a rapid in-house quality control test.

Highlights

Alpha-galactosidase (α-Gal) is an enzyme responsible for the hydrolyzation of glycolipids and glycoprotein commonly found in dietary sources
A typical example of a commercial α-galactosidase is that derived from Aspergillus niger mold, which is able to enact its function in the gastrointestinal tract by breaking down specific non-absorbable oligosaccharides before they are metabolized by colonic bacteria [15,17,18,19,22,23,24,25]
The enzymatic method was validated in the conditions established previously with the Plackett–Burman test, following both the ICH and the Food and Drug Administration (FDA) guidelines

Summary

Introduction

Alpha-galactosidase (α-Gal) is an enzyme responsible for the hydrolyzation of glycolipids and glycoprotein commonly found in dietary sources. Α-galactosidase is able to hydrolyze galacto-oligosaccharides such as starch, raffinose, melibiose and branched polysaccharides, galactomannans, and galacto-glucomannans, which catalyze the hydrolysis of α-1,6 linked galactose residues [1,2,3,4] This enzyme is produced in the mouth in the form of saliva, as well as in the pancreas from where it is moved into the small intestine and the rest of the digestive tract in order to perform its function [2,4]. To improve the nutritional value of products and make them digestible, α-galactosidases are often supplemented in food [12,13,14] These enzymes must resist various gut proteases in order to function properly in the human gut [2,4,6,7,15]. A typical example of a commercial α-galactosidase is that derived from Aspergillus niger mold, which is able to enact its function in the gastrointestinal tract by breaking down specific non-absorbable oligosaccharides before they are metabolized by colonic bacteria [15,17,18,19,22,23,24,25]

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