Rapid detection of methylglyoxal in blood samples

DESCRIPTION (provided by applicant): Diabetes is a serious, life-threatening disease caused by improper absorption and metabolism of glucose. Increased glucose levels can lead to an accumulation of toxic sugar degradation products, such as methylglyoxal (MG), in the bloodstream. MG is a reactive molecule which can modify and inactivate blood proteins which in turn leads to hypertension, neuropathy, and heart disease. Consequently, MG has been shown to be an excellent biomarker for a number of diabetes-related complications. Unfortunately, all current methods to directly measure MG levels in blood are very expensive, time-consuming and laborious. To fully realize the diagnostic potential of MG, rapid analytical methods are needed for the routine determinationof MG in biomedical samples such as blood. Our proposed Phase I research will create a unique test to measure MG levels in blood samples such as plasma and serum. This unique test will permit researchers to conveniently study the accumulation of reactiveMG in blood before damage to proteins and vasculature has occurred. Our novel assay will use a recombinant bacterial oxidoreductase enzyme (MG-R) capable of specifically and rapidly converting MG into a detectable colorimetric signal in a 96 well plate assay format. To boost sensitivity, we will link our enzymatic detection reaction to an NADP+/NADPH visible colorimetric amplification loop to increase assay sensitivity more than 100-fold. The assay will be optimized and validated using rat and mouse plasmaand serum spiked with known amounts of MG. After validation, we will commercially launch a rapid test kit to directly detect MG in preclinical and other biomedical samples. PUBLIC HEALTH RELEVANCE PUBLIC HEALTH RELEVANCE: Diabetes is a major health threat in the US; this life-threatening disease is rapidly spreading throughout our country (23 million Americans currently affected). Recently, scientists have discovered that a sugar metabolite called methylglyoxal plays a central role in the progression of diabetes - unfortunately there are no methods currently available for routine analysis of methylglyoxal. Our research will create a unique method to rapidly measure methylglyoxal levels in normal and diabetic blood samples; our new analytical method will provide researchers with a critical new diagnostic tool to more-effectively combat diabetes and associated cardiovascular disease.