Development of an equipment free paper based fluorimetric method for the selective determination of histidine in human urine samples.

A direct fluorimetric method, employing μicro-analytical paper-based devices (μ-PADs) for the selective determination of histidine (HIS) is described. The suggested method exploits the fluorescence emission of histidine after its rapid reaction with o-phthalaldehyde (OPA) at a basic medium (pH = 10) on the surface of a paper device with the application of a UV lamp at 354 nm. The devices are inexpensive and are composed of chromatographic paper and wax barriers. The analytical protocol is easily applicable with minimal technical expertise and without the need of expensive experimental apparatus. The user has to add a test sample, illuminate the device with a UV lamp, and read the fluorescence of the sensing area using a simple imaging device such as a cell-phone camera. The method is free from common interferences likely to affect the measurement of histidine and is selective among all other amino acids. This analytical procedure was optimized and validated, paying special attention to its intended application. The detection limits are as low as 1.8 μM with very satisfactory precision ranging from 6.4% (intra-day) to 8.9% (inter-day). Random urine samples from adult volunteers (n = 5) were successfully analyzed and HIS content ranged between 260 and 1114 μmol L with percentage recoveries in the range of 78.2 and 124.6%.