| Glycoproteins and glycolipids compounds are the most important parts of glycoconjugates. Glycoprotein, which is necessary for the body, has excellent physiological functions, and plays an important role in intramolecular communication in biological process, especially in cell-cell recognition, signal conditioning, cell differentiation and immune responses. However, it was found that sugar chains of glycoprotein exist microheterogeneity, and its content is limited in vivo. In addition, the synthesis process of glycoprotein is difficult. Therefore, people manage to find glycopeptide mimetics instead of glycoproteins during experimental study. Glycopeptide mimetics is a compound which is formed by a sugar conjugating with a specific amino acid or polypeptide by covalent bond. It retains the core of glycoprotein:the region of carbohydrate-peptide linker. Compared with glycoproteins, glycopeptides have a small molecular, simple structure, and are important models of glycoproteins. Meanwhile, during the synthesis process of the glycopeptide mimetics, glucosamine(2-amino-2-deoxy-D-glucose) is often used as glycosyl donor due to a wide range of raw materials resource and its biological activity.Firstly, we studied the synthesis of 1,3,4,6-four-O-acetylalpha-D-glucosaminesulfate(TAG). Then, made TAG and BTC react, generating isocyanate intermediates, respectively react with L-methionine methyl ester and L-isoleucine methyl ester, thus the target compound was synthesized. The purity of the two target products were 96.96% and 95.56% as detected by HPLC(Dionex). The molecular weights of the two products were 536 and 518 Da respectively as determined by ESI-MS mass spectrometry. The structure of the two products were confirmed by infrared spectroscopy, 1H NMRand 13 C NMR. The melting points of them were determined as 169~171.5℃ and 142~145℃.At last, we studied the anti-inflammatory effects of TAG-Ile and TAG-Met on RAW264.7 macrophages which inflammated by lipopolysaccharide. The results showed that TAG-Ile can control the release of IL-6 and NO, exerting its anti-inflammatory effect while TAG-Met had an obvious cell proliferation inhibition effect.The present study further intended to establish a method to quantitatively analyze glucosamine peptide mimetics with HPLC(Waters alliance 2695 and Dionex). Samples were separated by a Hypersil ODS2 column(5μm, 4.6 mm×250 mm), the mobile phase consisted of 5 mmol/L potassium dihydrogen phosphate-acetonitrile(p H=7.5). The flow rate was 1.0 m L/min, detection wavelength was 200 nm and column temperature was 20℃. The standard curve of four samples were linear over the range in 25.00~800.00 μg/m L, 40.00~1000.00 μg/m L, 27.00~600.00 μg/m L and 50.00~600.00 μg/m L, respectively. The limit of quantitation were 20.00, 35.54, 25.00 and 49.00 μg/m L in turn(S/N=10). Limit of detection were 4.50, 15.30, 13.45 and 19.30 μg/m L(S/N=3). The average recovery rate of the four samples were 99.00~101.42%, day and inter-day precision were less than 1.37%. The method is simple, rapid, specific, and suitable for the determination of glucosamine peptides. |
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