In vitro human serum albumin glycation monitored by Terahertz spectroscopy
Maria Mernea, Alina Ionescu, Ionut Vasile, Cristina Nica, Gheorghe Stoian, Traian Dascalu, Dan Florin Mihailescu
The nonenzymatic attachment of sugars to proteins, namely glycation, is accelerated under diabetic conditions. Monitoring the glycated human serum albumin (HSA) levels gives the short term variation of glucose concentration in diabetic patients blood. Therefore, a significant effort was made to measure glycated HSA, including by spectroscopic methods such as Raman. Here we used THzspectroscopy to monitor HSA glycation in time (over 5, 7 and 11 weeks). Different sugar types have different reactivity; therefore we also addressed the reducing sugar influence on glycation by performing in vitro HSA glycation by both glucose and fructose. Since residues protonation state influences their susceptibility for glycation, we incubated HSA with sugars at two pH values: 7 and 8. Our results show that THz absorption decreases with the incubation time of HSA with sugars. At the incubation times we considered, the most significant differences were obtained on HSA samples glycated using glucose. Differences between samples glycated by glucose and by fructose show that glycation by glucose is a slower process. At pH 7, glycation by glucose is slower than at pH 8, while glycation by fructose is slightly faster at pH 7 than at pH 8. Glycated HSA models with different degrees of glycation were built by molecular modeling. Simulated THz spectra of the models are in good agreement with the experimental data. All these show that THz spectroscopy could monitor the progression of glycation in time and that it is sensitive to reducing sugars or pH value used in the glycation process.