Department of Medicinal Chemistry, Faculty of Pharmacy, University of El Mounufia, El Mounufia, Egypt.
Department of Analytical Chemistry, Faculty of Pharmacy, Tanta University, Tanta, Egypt.
Luminescence. 2020 Mar;35(2):284-291. doi: 10.1002/bio.3725. Epub 2019 Nov 24.
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 with a lower detection limit of 0.29 ng ml and a lower quantification limit of 0.88 ng ml . 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.
尚未实现使用荧光直接测定苯甲酸阿格列汀（ALO），因为 ALO 不能自然荧光。因此，应首先用荧光衍生试剂对其进行衍生化，以提高生物分析所需的灵敏度。本方法是利用其氨基的亲核性质，在硼酸盐缓冲液 pH 8.5 下与 4-氯-7-硝基苯并呋咱（NBD-Cl）反应，生成强荧光化合物，然后在 470nm 和 527nm 处分别激发和发射，首次建立了用于 ALO 定量的荧光分光光度法。仔细研究并优化了反应和荧光强度的实验变量。线性范围为 1-250ngml，检测限为 0.29ngml，定量限为 0.88ngml。通过片剂中平均回收率为 100.62±1.59%，人血浆中为 99.86±0.82%，验证了本研究的准确性。此外，该方法已用于口服给予 ALO 后大鼠真实血浆中的生物分析，样品制备简单。该方法已被证明是生物等效性研究中 ALO 分析的一种很有前途的替代方法。
