Hollman P C, van Trijp J M, Buysman M N
DLO [Formula: see text] State Institute for Quality Control of Agricultural Products (RIKILT-DLO), Bornsesteeg 45, 6708 PD Wageningen, The Netherlands.
Anal Chem. 1996 Oct 1;68(19):3511-5. doi: 10.1021/ac960461w.
Flavonols are dietary antioxidants which may prevent coronary heart disease. To be able to study absorption of flavonols in humans, we developed a postcolumn derivatization with aluminum for HPLC with fluorescence detection. Variables governing postcolumn chelation, such as water content, buffer, organic modifier of the eluent, concentration of Al(3+), presence of acetic acid in the postcolumn reagent, and temperature, were studied and optimized. Of the flavonoids, only flavonols that contain a free 3-hydroxyl and 4-keto oxygen binding site form fluorescent complexes with Al(3+). The method has a detection limit of 0.15 ng/mL for quercetin, 0.05 ng/mL for kaempferol, 0.45 ng/mL for myricetin, and 0.05 mg/mL for isorhamnetin, thus improving detectability of quercetin 300-fold as compared to that possible with UV detection. The reproducibility relative standard deviation of the method is 1.4%. This extremely sensitive method enables, for the first time, determination of flavonols in body fluids after consumption of a normal diet.
黄酮醇是膳食抗氧化剂，可能预防冠心病。为了能够研究人体对黄酮醇的吸收情况，我们开发了一种用于高效液相色谱荧光检测的铝柱后衍生化方法。研究并优化了柱后螯合的相关变量，如水含量、缓冲液、洗脱液的有机改性剂、Al(3+)浓度、柱后试剂中乙酸的存在以及温度。在黄酮类化合物中，只有含有游离3-羟基和4-酮氧结合位点的黄酮醇能与Al(3+)形成荧光络合物。该方法对槲皮素的检测限为0.15 ng/mL，对山奈酚为0.05 ng/mL，对杨梅素为0.45 ng/mL，对异鼠李素为0.05 mg/mL，因此与紫外检测相比，槲皮素的可检测性提高了300倍。该方法的重现性相对标准偏差为1.4%。这种极其灵敏的方法首次能够测定正常饮食后体液中的黄酮醇。
