Instrumentation and techniques for analysis of hydrogen peroxide and peroxide-producing reactions involving earthworm (Diplocardia longa) bioluminescence.

Earthworm bioluminescence (species Diplocardia longa) is a sensitive tool for determination of peroxide titers in biological systems, with linearity of the standard curve for hydrogen peroxide extending from 10 nmol/L in the assay to greater than 1 mmol/L, and with sensitivity to 2 pmol of peroxide per assay. This assay's insensitivity to extraneous protein, turbidity, and solutes that are not redox active makes it particularly useful for automated or crude sample analysis and for analysis of oxidases and their substrates in coupled reactions. Examples of this latter application we discuss are analyses of glucose and putrescine oxidases and their substrates. In each case linear standard curves are obtained, covering several decades of concentration. The coupled reactions also allow in situ detection of oxidases in electrophoresis gels. We also describe a small, portable photometer system and an on-line computer-controlled instrument system that has facilitated development of these assays and will also allow automatic scanning of bioluminescence in gels. The data-collection and decision-making algorithms of this system are simple and easily adaptable for use in an inexpensive computer-based photometer system.