The direct determination of alogliptin benzoate (ALO) using fluorescence has not yet been accomplished because ALO cannot fluoresce naturally. Accordingly, it should be derivatized first with a fluorogenic reagent to enhance the sensitivity required for its bioanalysis. This method is the first spectrofluorimetric assay for ALO quantification exploiting the nucleophilic nature of its amino group to react with 4-chloro-7-nitrobenzofurazan (NBD-Cl) in borate buffer at pH 8.5 to produce a strong fluorescent compound that is excited at and emits at wavelengths 470 and 527 nm, respectively. Experimental variables concerning the conditions of reaction and fluorogenic intensity were carefully investigated and optimized. Linearity was from 1-250 ng ml-1 with a lower detection limit of 0.29 ng ml-1 and a lower quantification limit of 0.88 ng ml-1 . Validation of the current study was accomplished with mean per cent recovery of 100.62 ± 1.59 in tablets and 99.86 ± 0.82 in human plasma. Furthermore, the current method has been utilized in the bioanalysis of ALO in real rat plasma after oral administration with a simple specimen preparation. The developed method has proven to be a promising alternative method for ALO analysis in bioequivalence studies.

中文翻译：

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利用其与4-氯-7-硝基苯并呋喃的相互作用，对生物流体中的苯甲酸阿格列汀进行了新的荧光光谱定量分析。

由于ALO不能自然发出荧光，因此尚未完成使用荧光法直接测定苯甲酸阿格列汀（ALO）的方法。因此，应先用荧光试剂将其衍生化，以增强其生物分析所需的灵敏度。该方法是第一种用于ALO定量的荧光光谱测定法，利用其氨基的亲核性质与pH 8.5的硼酸盐缓冲液中的4-氯-7-硝基苯并呋喃（NBD-Cl）反应生成强荧光化合物，该化合物在和分别以470和527 nm的波长发射。仔细研究和优化了有关反应条件和荧光强度的实验变量。线性范围为1-250 ng ml-1，检测下限为0.29 ng ml-1，定量下限为0.88 ng ml-1。当前研究的验证是通过片剂中100.62±1.59的平均回收率和人血浆中99.86±0.82的平均回收率来完成的。此外，在通过简单的样品制备物口服给药后，目前的方法已用于真实大鼠血浆中ALO的生物分析。实践证明，开发的方法是生物等效性研究中ALO分析的有前途的替代方法。