Department of Chemistry, The Islamia University of Bahawalpur, Bahawalpur 63100, Pakistan.
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, The Islamia University of Bahawalpur, Bahawalpur 63100, Pakistan.
Bioorg Chem. 2021 May;110:104818. doi: 10.1016/j.bioorg.2021.104818. Epub 2021 Mar 10.
Here we report the inhibitory effects of nine non-steroidal anti-inflammatory drugs (NSAIDs) on soybean 15-lipoxygenase (15-LOX) enzyme (EC 1.13.11.12) by three different methods; UV-absorbance, colorimetric and chemiluminescence methods. Only two drugs, Ibuprofen and Ketoprofen, exhibited enzyme inhibition by UV-absorbance method but none of the drug showed inhibition through colorimetric method. Chemiluminescence method was found highly sensitive for the identification of 15-LOX inhibitors and it was more sensitive and several fold faster than the other methods. All tested drugs showed 15-LOX-inhibition with IC values ranging from 3.52 ± 0.08 to 62.6 ± 2.15 µM by chemiluminescence method. Naproxen was the most active inhibitor (IC 3.52 ± 0.08 µM) followed by Aspirin (IC 4.62 ± 0.11 µM) and Acetaminophen (IC 6.52 ± 0.14 µM). Ketoprofen, Diclofenac and Mefenamic acid showed moderate inhibitory profiles (IC 24.8 ± 0.24 to 39.62 ± 0.27 µM). Piroxicam and Tenoxicam were the least active inhibitors with IC values of 62.6 ± 2.15 µM and 49.5 ± 1.13 µM, respectively. These findings are supported by expression analysis, molecular docking studies and density functional theory calculations. The expression analysis and flow cytometry apoptosis analysis were carried out using mononuclear cells (MNCs) which express both human 15-LOX and 5-LOX. Selected NSAIDs did not affect the cytotoxic activity of MNCs at IC concentrations and the cell death showed dose dependent effect. However, MNCs apoptosis increased only at the higher concentrations, demonstrating that these drugs may not induce loss of immunity in septic and other inflammatory conditions at the acceptable inhibitory concentrations. The data collectively suggests that NSAIDs not only inhibit COX enzymes as reported in the literature but soybean 15-LOX and MNCs LOXs are also inhibited at differential values. A comparison of the metabolomics studies of arachidonic acid pathway after inhibition of either COX or LOX enzymes may reconfirm these findings.
在这里，我们通过三种不同的方法，即紫外吸收法、比色法和化学发光法，报告了九种非甾体抗炎药（NSAIDs）对大豆 15-脂氧合酶（15-LOX）酶（EC 1.13.11.12）的抑制作用。只有两种药物，布洛芬和酮洛芬，通过紫外吸收法显示出酶抑制作用，但没有一种药物通过比色法显示出抑制作用。化学发光法被发现对 15-LOX 抑制剂的鉴定非常敏感，它比其他方法更敏感，速度也快几倍。通过化学发光法，所有测试的药物均显示出对 15-LOX 的抑制作用，其 IC 值范围为 3.52±0.08 至 62.6±2.15µM。萘普生是最有效的抑制剂（IC 3.52±0.08µM），其次是阿司匹林（IC 4.62±0.11µM）和对乙酰氨基酚（IC 6.52±0.14µM）。酮洛芬、双氯芬酸和甲芬那酸显示出中等抑制谱（IC 24.8±0.24 至 39.62±0.27µM）。吡罗昔康和替诺昔康是最不活跃的抑制剂，IC 值分别为 62.6±2.15µM 和 49.5±1.13µM。这些发现得到了表达分析、分子对接研究和密度泛函理论计算的支持。使用单核细胞（MNCs）进行表达分析和流式细胞术凋亡分析，单核细胞表达人 15-LOX 和 5-LOX。所选 NSAIDs 以 IC 浓度不会影响 MNCs 的细胞毒性活性，并且细胞死亡显示出剂量依赖性效应。然而，只有在较高浓度下，MNCs 的凋亡才会增加，这表明在可接受的抑制浓度下，这些药物不会在脓毒症和其他炎症情况下引起免疫丧失。数据表明，NSAIDs 不仅如文献中报道的那样抑制 COX 酶，而且还以不同的数值抑制大豆 15-LOX 和 MNCs LOX。抑制 COX 或 LOX 酶后对花生四烯酸途径的代谢组学研究的比较可能会再次证实这些发现。
