新型经皮传递的外周脂肪细胞激动剂衍生物TAT-hNPY的实验研究及其临床意义

An Experimental Study of a New Transdermal Peripheral Fat Cell Agonist Derivative and Its Clinical Significance

目的为了未来开发和研制一类具有经皮肤或经粘膜途径给药的新型外周脂肪细胞激动剂及进行相关临床应用制剂等研究工作需要,本研究新设计并合成了一条氨基酸全序列为YGRKKRRQRRRYPSKPDNPGEDAPAEDMARYYSALRHYINLITRQRY-NH2的TAT-hNPY,并对TAT-hNPY的纯度和质核比及分子量进行了测定分析与鉴定,为其在临床应用提供实验依据。方法 TAT-hNPY合成采用化学固相合成法,纯度测定采用高效液相色谱法(HPLC),分子量测定采用电喷雾离子化质谱法(ESI-IT-MS)。结果采用HPLC检测化学固相法合成的。TAT-hNPY的纯度为95.85%;采用ESI-IT-MS法检测分析出3个带有不同质荷比的目标峰,它们分别为:[M+6H]6H+质荷比为969.2 m/z,其实测分子量为5 809.2道尔顿;[M+7H]7H+质荷比为:831.4m/z,其实测分子量为5 812.8道尔顿;[M+9H]9H+质荷比为:646.8m/z,其实测分子量为5812.2道尔顿,该目标多肽的理论分子量为5814.6道尔顿。结论采用化学固相合成法合成TAThNPY的实测分子量与理论分子量误差均在允许误差0.1%内,两者结果相符,证明本实验成功实现了目标多肽的合成,这将为临床相关学科进一步探索hNPY与外周脂肪细胞之间的相关关系、特别是对深入探索研究经皮肤或粘膜给药途径传递hNPY对人体外周组织脂肪组织中脂肪前体细胞和脂肪干细胞的相互作用及其影响等奠定了一定基础。

Objective To measure and identify the purity, mass-to-charge ratio（m/e） and molecular weight of the synthesized TAT-hNPY with YGRKKRRQRRRYPSKPDNPGEDAPAEDMARYYSALRHYI-NLITRQRY-NH2 as its full amino acid sequence, a new transdermal peripheral fat crell agonist derivative so as to provide basis for clinical application. Methods Chemical solid phase synthesis was adopted for synthesis of TAT-hNPY, high performance liquid chromatography（HPLC）was applied in purity determination, and electrospray ionization-mass spectrometry（ESI-MS） in molecular weight measurement.Results The HPLC detection indicated that the purity of TAT-hNPY produced by chemical solid phase synthesis reached 95.85%;ESI-MS measurement revealed 3 target peaks with different charges： the mass-to-charge ratio of [M＋6H] 6H＋ of 969.2 m/z with the measured molecular weight of 5 809.2 Dalton, the mass-to-charge ratio of [M＋7H] 7H＋ of 831.4 m/z with the measured molecular weight of 5 812.8 Dalton and the mass-to-charge ratio of [M＋9H] 9H＋ of 646.8 m/z with the measured molecular weight of 5 812.2 Dalton; the theoretical molecular weight was 5 814.6 Dalton. Conclusions The variation between the measured molecular weight of TAT-hNPY synthesized by chemical solid phase synthesis and the theoretical molecular weight is less than the acceptable error range of 0.1%, the consistence proves the success in the design and synthesis of the target polypeptide, which provides basis for further exploration of the correlation between hNPY and peripheral fat cells, especially for further exploration of the effect of hNPY delivered through skin or mucosal upon fat precursor cells and stem cells of human peripheral fat tissue.