A modified enzymatic method for measuring insect sugars and the effect of storing samples in ethanol on subsequent trehalose measurements

A modified method for enzymatically measuring concentrations of glucose, fructose and trehalose in parasitoid wasps is described and evaluated. The method has high specificity, gives results comparable to high performance liquid chromatography (HPLC), and shows good correlations with HPLC. The enzymatic method is quicker and less expensive than HPLC, and is safer, faster and more sensitive than another commonly used insect sugar determination method, the anthrone test. The method has potential to measure additional sugars, and details for measuring sucrose are provided as Supplementary material. We also investigate if endogenous enzymes can retain activity in insects preserved in ethanol, thus potentially skewing insect sugar measurements. The results show that the enzyme trehalase, which converts the insect haemolymph sugar trehalose to glucose, can remain active in insect specimens that are stored whole in ethanol. Thus, sugar measurements from insects preserved in this way have potential for bias towards low trehalose and high glucose. In contrast, the activity of endogenous trehalase in insects that are either frozen at −20 °C, or crushed in ethanol is either much reduced or eliminated. If, prior to sugar analysis, insects are to be stored in ethanol at temperatures above −20 °C, then we recommend they be crushed.