Abstract
A novel method for quantitation of cardiac muscle carnosine levels using HPLC-UV is described. In this simple and reliable method, carnosine from the rat cardiac muscle and the internal standard, thymopentin, were extracted by protein precipitation with acetonitrile. The method was linear up to 60.96 μg·mL−1 for L-carnosine. The calibration curve was linear in concentration ranges from 0.5 to 60.96 μg·mL−1. The relative standard deviations obtained for intra- and interday precision were lower than 12% and the recoveries were higher than 90% for both carnosine and internal standard. We successfully applied this method to the analysis of endogenous carnosine in cardiac muscle of the diabetes rats and healthy control rats. The concentration of carnosine was significantly lower in the diabetes rats group, compared to that in the healthy control rats. These results support the usefulness of this method as a means of quantitating carnosine and illustrate the important role of L-carnosine in cardiac muscle.
Highlights
Carnosine (Figure 1(a)), N-beta-alanyl-L-histidine, an endogenous material, has been found in skeletal muscle [1, 2], brain, olfactory bulbs [3], and crystalline lens [4, 5]
The calibration curve of carnosine was at concentrations of 0.51, 1.02, 2.54, 10.16, 20.32, 40.64, and 60.96 μg⋅mL−1 which were prepared by spiking appropriate amount of the standard solution in diluted blank cardiac muscle homogenates
In view of the liquid-liquid extraction usually offering much cleaner sample, various organic reagents have been tried for liquid-liquid extraction method but the great polarity of carnosine should lead to the failure
Summary
Carnosine (Figure 1(a)), N-beta-alanyl-L-histidine, an endogenous material, has been found in skeletal muscle [1, 2], brain, olfactory bulbs [3], and crystalline lens [4, 5]. The current methods for measuring carnosine in biologic matrices include micellar liquid chromatography [2], high-performance anion-exchange chromatography [16, 17], HPLC-MS [18], and HPLC-UV with precolumn derivatization [19, 20] and without derivatization using NH2 column [1]. All these methods suffer from several drawbacks: tedious micellar mobile phases preparation and care of the column procedure, instability of derivatives, and the high cost of MS detector preventing their utilization by many laboratories. We successfully applied this method to the analysis of carnosine in the diabetes rat cardiac muscle
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