青光眼家兔玻璃体内游离谷氨酸盐浓度的测定

Determination of free glutamate concentration in vitreous humor of rabbits with glaucoma

关键词:  高效液相荧光检测法  葛根素和灯盏乙素阳离子脂质纳米粒  青光眼模型  玻璃体游离谷氨酸盐

Key words:  high performance liquid phase fluorescence detection  Puerarin and Scutellarin cationic lipid nanoparticles  glaucoma model  vitreous humor free glutamate

摘要:

[目的]建立高效液相荧光检测法，测定青光眼家兔玻璃体游离谷氨酸盐浓度。[方法]于青光眼家兔玻璃体分别灌注葛根素和灯盏乙素阳离子脂质纳米粒和药物溶液，在色谱条件为Diamonsil C₁₈色谱柱（200 mm×4.6 mm，5 μm），流动相为50 mmol磷酸盐缓冲液：甲醇（25：75），流速为1.0 mL/min，柱温30℃，荧光检测器的激发波长（λex）350 nm，发射波长（λem）450 nm下测定透析液中游离谷氨酸盐浓度。[结果]玻璃体内植入探针的在体回收率为33.60%（r=0.970 6）。在玻璃体内灌注葛根素和灯盏乙素阳离子脂质纳米粒组与药物溶液组后，测定不同时间点的游离谷氨酸盐浓度，结果表明与溶液组相比，制剂组谷氨酸盐浓度降低1.59倍，有统计学差异（P<0.05）。[结论]该方法简便、准确、可靠，为玻璃体游离谷氨酸盐的测定提供参考。

Abstract:

[Objective] To establish a high performance liquid phase fluorescence assay for the determination of free glutamate concentration in vitreous humor of rabbits with glaucoma.[Methods] Puerarin and Scutellarin cationic lipid nanoparticles (PUE-SCU CLN) and drug solution were injected into the vitreous humor of glaucoma rabbits. The chromatographic conditions were Diamonsil C₁₈ column (200 mm×4.6 mm, 5 μm), the excitation wavelength (λex) of the fluorescence detector is 350 nm, and the emission wavelength (λem) is 450 nm, the mobile phase consisted of 50 mmol phosphate buffer:methanol (25:75) at a flow rate of l.0 mL/min and a columnar temperature of 30℃. The concentration of free glutamate in the dialysate was measured.[Results] The in vivo recovery of vitreous humor implant was 33.60% (r=0.970 6). The concentration of free glutamate at different time points was measured after vitreous humor injection of PUE-SCU CLN and drug solution. The results showed that the concentration of glutamate in the preparation group was lower than that in the solution group 1.59 times, there was significant difference (P<0.05).[Conclusion] The method is simple, accurate and reliable, and it can be used as a reference for the determination of free glutamate in vitreous humor.