Faculty of Chemistry, Department of Environmental Chemistry, University of Lodz, 163 Pomorska Str., 90-236 Łódź, Poland.
J Agric Food Chem. 2020 Sep 30;68(39):10992-10999. doi: 10.1021/acs.jafc.0c03778. Epub 2020 Sep 19.
During this research a simple, accurate, and environmentally friendly method to determine lipoyllysine and lipoic acid in meat was developed and validated. The presented approach was based on the hydrolysis of the proteins containing lipoic acid, reduction of disulfide bonds with tris(hydroxymethyl)phosphine, and precolumn derivatization of free thiol groups with 1-benzyl-2-chloropyridinium bromide long-term followed by HPLC separation with a diode-array detector. The method has been validated in accordance with the U.S. FDA guidelines and was linear in the range of 0.1-10 μmol/L in concentration with values ≥0.9997 for both analytes. For lipoyllysine and lipoic acid, intra- and interday precision values were lower than 10%. The intraday accuracy values ranged from 91.0% to 99.4% for lipoyllysine and from 99.1% to 107.3% for lipoic acid, whereas the interday accuracy values for lipoyllysine and lipoic acid were 92.0-95.6% and 93.5-98.8%, respectively. Additionally, in this research the antioxidant activity of lipoyllysine and reduced lipoyllysine compound using spectrophotometric method with 1,1-diphenyl-2-picrylhydrazyl was examined for the first time. The data showed that dihydrolipoyllysine exhibits stronger antioxidant capacity than lipoyllysine based on a lower value of concentration required to achieve a 50% antioxidant effect in 1,1-diphenyl-2-picrylhydrazyl radical scavenging test.
在这项研究中，开发并验证了一种简单、准确且环保的方法来测定肉中的二氢硫辛酸和游离硫辛酸。所提出的方法基于蛋白质中脂酰赖氨酸的水解、二硫键与三（羟甲基）膦的还原，以及游离巯基与 1-苄基-2-氯吡啶溴化物的预柱衍生化，随后进行高效液相色谱分离和二极管阵列检测。该方法符合美国 FDA 指南的要求，在 0.1-10 μmol/L 的浓度范围内具有良好的线性关系，两个分析物的 r 值均≥0.9997。对于二氢硫辛酸和游离硫辛酸，日内和日间精密度值均低于 10%。二氢硫辛酸的日内准确度值范围为 91.0%-99.4%，游离硫辛酸的准确度值范围为 99.1%-107.3%，而二氢硫辛酸和游离硫辛酸的日间准确度值分别为 92.0%-95.6%和 93.5%-98.8%。此外，本研究首次采用分光光度法，以 1,1-二苯基-2-苦基肼基（DPPH）为自由基清除剂，研究了二氢硫辛酸和还原型二氢硫辛酸化合物的抗氧化活性。数据表明，二氢硫辛酸在 DPPH 自由基清除试验中，达到 50%抗氧化效果所需的浓度值低于二氢硫辛酸，表明其具有更强的抗氧化能力。
