Determination Of Several Small Molecule Compounds In Complex Matrix By Dispersive Liquid-liquid Extraction Combined With High Phase Liquid Chromatograph

Dispersive liquid liquid microextraction(DLLME) is a new microextraction and environment friendly pretreatment method, which reported by Rezaee in 2006. It is simple, convenient, saves time and easy to operate, has excellent precision and good selectivity and owns multi-element enrichment capability and low detection limits. DLLME has a good compatibility characteristic to combine with high phase liquid chromatography(HPLC), high phase liquid chromatography-mass spectrometry(HPLC-MS), gas chromatography(GC), gas chromatography-mass spectrometry(GC-MS) and so on. It is applied widely, including environmental pollutants control, analytical chemistry,biochemistry, clinical medicine, pharmacological researches. In this paper,a method on dispersive liquid-liquid microextraction for separation and analysis of several small molecule compounds in complex matrix prior to determination by high performance liquid chromatography has been studied.It mainly consists of the following four parts:1. The summary of several sample pretreatment methods, especially liquid phase microextraction(LPME) and dispersed liquid liquid phase microextraction(DLLME).2. A simple and fast carrier-mediated dispersive liquid-liquid microextraction method followed by high performance liquid chromatography(HPLC) was developed to selectively extract and measure urinary monoamine neurotransmitter metabolites(MNMs). Weoptimized factors influencing extraction such as extraction and dispersant solvent type and volume, sample solution pH, carrier concentration,extraction time. We used tributyl phosphate(TBP) as a carrier and n-butanol and methanol as extraction and dispersive solvents, respectively.Using optimal conditions, enrichment factors for 5-hydroxyindoleacetic acid(5-HIAA), vanillylmandelic acid(VMA) and homovanillic acid(HVA) were 34, 21 and 32, respectively. Linearity was investigated over a range of 5~1000 μg/L. Limits of detection(S/N = 3) were 2 μg/L for HVA and VMA, 1 μg/L for 5-HIAA. Spiked sample recovery were93.55%~108.65%, and RSD were less than 7.6%(n = 3). The method was successfully used to separate and enrich urinary MNMs.3. A novel method for the determination of vardenafil and sildenafil in complex samples by dispersive liquid-liquid microextraction-back extraction-solidification of acceptor phase(DLLME-BE-SAP)-high performance liquid chromatography was developed. The optimal experimental conditions were as follows: 200 μL toluene as the organic phase, 60 μL methanol as the dispersive solvent, the mixture of 19 μL of0.1 mol/L HCl and 1 μL methanol as the acceptor phase. The extraction time was 2 min. High enrichment factor(47-fold) of sildenafil and(46-fold) of vardenafil were obtained under the optimal experimental conditions. The linear range was 1~1000 μg/L. The limit of detection(S/N = 3) of 0.5 μg/L for sildenafil and 0.8 μg/L for vardenafil. The recoveries of the spiked samples were from 95.5% to 109.7% and the inter-day relative standard deviations were lower than 7.6%.The proposed method may avoid the pollution of the chromatographic column due to injecting directly with organic solvents, extend the range of available organic extraction solvents in DLLME based on solidification of floating organic drop, improve the compatibility of the method with HPLC analysis, and was found to be convenient, rapid, accurate, sensitive andenvironment friendly.4. A novel method for the determination of glipizide and gliclazide in dietary supplement by dispersive liquid-liquid microextraction back extraction solidification of acceptor phase(DLLME-BE-SAP)-high performance liquid chromatography was developed. The optimal experimental conditions were as follows: 200 μL toluene as the organic phase, 30 μL methanol as the dispersive solvent, the mixture of 14 μL of0.04 mol/L NaOH and 1 μL methanol as the acceptor phase. The extraction time was 90 sec. High enrichment factor(26-fold) of glipizide and(24-fold) of gliclazide were obtained under the optimal experimental conditions. The linear range was 1~1000 μg/L. The limit of detection(S/N = 3) of 1 μg/L for glipizide and 2 μg/L for gliclazide. The recoveries of the spiked samples were from 91.8% to 107.6%, and the inter-day relative standard deviations were lower than 9%.The method has provided high analyte preconcentration and is a sensitive and suitable method for the simultaneous determination of glipizide and gliclazide in dietary supplements.