Removing small non-enzymatic molecules for biochemical assay of redox regulatory enzymes; An exemplary comments on "Antioxidant responses in gills and digestive gland of oyster Crassostrea madrasensis (Preston) under lead exposure

Abstract

For biochemical assay of every enzyme including redox regulatory enzymes, any interfering small molecules (ISM) that may cross react with substrates or indirectly influence the reaction, must be removed. Such ISM(s) if present, need to be either neutralized or filtered out both from the sample or from the reaction mixture. This is a standard protocol adapted worldwide and fundamental rule in biochemistry. Without such approach, results obtained from a study that includes enzymatic assays seem to be inaccurate. Such inaccuracy raises question on the use of such data in future, especially as ecotoxic biomarkers. Tissue specific seasonal variation in natural titre of such ISM(s) in organisms leads to give rise counterfeit results. Such a case is highlighted in this correspondence article in relation to assay of redox regulatory enzymes including superoxide dismutase, catalase, especially the enzymes of glutathione system in presence of glutathione (GSH) in sample. This fact is discussed considering a recent publication doi.org/10.1016/j.ecoenv.2017.03.056 in which the authors have measured glutathione enzymes in tissues without removing GSH, that acts as ISM, from the sample. It is inferred that logical and sound scientific practices need to be followed for measuring biochemical activity of all enzymes in general and enzymes of glutathione system in particular. The main objective of this article was to make an alert in scientific society to avoid such mistakes in future.

Keywords: Ecotoxic biomarker; Glutathione; Glutathione peroxidase; Glutathione reductase; Inaccurate method; Superoxide dismutase.