Hemoglobin/Myoglobin/Creatinine Urine Detection

Rhabdomyolysis is a dangerous disorder that can occur after a crush (compression) injury to muscle tissue in the human body. Clinicians want to be able to detect myoglobin, above a certain level, which the affected muscle tissue releases into the urine stream and blood stream. The myoglobin must then be distinguished from hemoglobin, which also may be released into the urine stream due to a rupture of red blood cells from a crush injury. Myoglobin present in the bloodstream presents immediate danger of kidney failure while hemoglobin is a lesser problem. The current method to detect hemoglobin and myoglobin in urine is a dipstick test. This can alert the clinician to the heme present in urine, but heme is a substance that is present in both hemoglobin and myoglobin molecules.

Using Raman Spectrescopy for detection and quantification of these chemicals has diagnosis potential here. Raman Spectroscopy can detect the two different spectra, thus indicating the chemicals presence, and also be able to determine concentrations by relative intensities of peaks in the spectra. Raman spectroscopy also allows for a simultaneous measurement of creatinine, a byproduct in urine that is released in constant amounts by the kidneys. By measuring the concentrations of creatinine, the concentrations of hemoglobin and myoglobin may be normalized to the varying water content in urine.

Raman Spectra of highly concentrated hemoglobin and myoglobin exhibit some differences in vibrational energies (shown below). These differences are attributed to differences in protein chains surrounding the central iron containing heme unit, while the similarities are attributed to the heme unit itself which has strongest raman bands in the molecule. Heme's vibrational bands are widely studied and may be precisely assigned through the literature. Experimentation with simulated urine data, i.e. patient urine doped with analyte levels above physiological values, has proved to be quite successful. Prediction plots of predicted and doped concentrations for a typical experiment are shown below. Further experimentation has shown promise in detection of native creatinine and good potential for the ultimate goal of detection of native hemoglobin and myoglobin.