Molecular docking, molecular dynamics, MM/PBSA approaches and bioactivity studies of nepetanudoside B isolated from endemic Nepeta aristata.

Nepetanudoside B (NNB) was isolated from aerial parts of endemic Nepeta aristata crude extract (CH3OH-CHCl3) using silica gel (n-hexane, methanol, ethyl acetate, and dichlorometane, respectively) and sephadex LH-20 (65% Methanol-35% Chloroform) column chromatographies. Preparative-HPLC was used to purify NNB after activity-guided isolation of methanol sub-fractions with enzyme inhibitory and DNA protective properties. The NNB was determined using 1H,13C, COSY, HSQC, HMBC, and LC-MS/MS. The study compared the effects of NNB with conventional drugs in terms of its ability to inhibit enzymes such as urease, α-amylase, carbonic anhydrase (CA), lipase, α-glucosidase, and tyrosinase, as well as its ability to protect DNA. Enzyme kinetic and molecular docking were also used to evaluate this. NNB exhibited the best inhibitory activity on urease (1.28 ± 0.00 µg/mL), lipase (5.83 ± 0.10 µg/mL), BChE (3.73 ± 0.46 µg/mL), tyrosinase (7.39 ± 0.00 µg/mL), α-glucosidase (10.95 ± 0.00 µg/mL), α-amylase (22.11 ± 1.03 µg/mL) and AChE (25.68 ± 3.32 µg/mL), respectively. NNB has higher MolDock scores with binding energy in α-glucosidase (-233) and BChE (-8.90 kcal/mol). In enzyme kinetics studies, it was determined that urease, AChE, α-glucosidase, lipase, and CA were non-competitive , while BChE and tyrosinase were competitive inhibition mechanisms. Their Ki values were calculated as 0.09, 0.24, 0.09, 0.10, 0.08, 0.05, and 0.07 mM, respectively. Molecular dynamics simulation studies were performed for the interactions of NNB-BChE with MM/PBSA binding free energey RMSD, RMSF, Rg, SASA, and also the number of hydrogen bonds was calculated. The suitability and effectiveness of NNB have been proven in the food and pharmaceutical industries. The NNB molecule may lead to development studies as a BChE inhibitor.Communicated by Ramaswamy H. Sarma.

DNA protection, molecular docking, enzyme inhibition and enzyme kinetic studies of 1,5,9-epideoxyloganic acid isolated from Nepeta aristata with bio-guided fractionation.

1,5,9-epideoxyloganic acid (ELA) was isolated from the aerial parts of endemic Nepeta aristata Boiss Et Kotschy Ex Boiss crude extract (methanol:chloroform) using silica gel (hexane, chloroform, ethyl acetate, and methanol) and sephadex LH-20 (65% methanol-35% chloroform) columns. Activity-guided isolation was performed on methanol sub-fractions with DNA protection and enzyme inhibitory activities, and then the ELA was purified by prep-HPLC. The ELA structure, bio-guided isolate, was determined via 1H NMR, 13C NMR, and MS spectrometry. ELA's enzyme inhibition and DNA protection activities were investigated and compared with standard drugs. The inhibition capacity of ELA against acetylcholinesterase (AChE), butyrylcholinesterase (BChE), urease, carbonic anhydrase (CA), α-glucosidase, α-amylase, lipase, and tyrosinase enzymes was evaluated by kinetic and molecular docking results. The ELA displayed the best inhibitory activity on AChE, BChE, α-glucosidase, urease, α-amylase, and tyrosinase with IC50 values of 2.53 ± 0.27, 3.75 ± 0.11, 3.98 ± 0.07, 4.40 ± 0.01, 6.43 ± 0.54 and 7.39 ± 0.00 µg/mL, respectively. ELA acted as a competitive inhibitor against BChE and α-glucosidase and a non-competitive inhibitor against AChE. The ELA's binding affinity values on AChE, BChE, and α-glucosidase were -7.70, -8.50, and -8.30 kcal/mol, respectively. DNA protection activity of the ELA molecule was determined as 57.53% for form I and 53.57% for form II. In conclusion, the inhibitory activity of ELA demonstrated its effectiveness in terms of its suitability in the pharmaceutical industry.Communicated by Ramaswamy H. Sarma.

Determination of antioxidant, DNA protection, enzyme inhibition potential and molecular docking studies of a biomarker ursolic acid in Nepeta species.

Ursolic acid (UA), which has many biological properties such as anti-cancer, anti-inflammatory and antioxidant, and regulates some pharmacological processes, has been isolated from the flowers, leaves, berries and fruits of many plant species. In this work, UA was purified from the methanol-chloroform crude extract of Nepeta species (N. aristata, N. baytopii, N. italica, N. trachonitica, N. stenantha) using a silica gel column with chloroform or ethyl acetate solvents via bioactivity-guided isolation. The most active sub-fractions were determined under bioactivities using antioxidant and DNA protection activities and enzyme inhibitions. UA was purified from these fractions and its structure was elucidated by NMR spectroscopy techniques. The highest amount of UA was found in N. stenantha (8.53 mg UA/g), while the lowest amount of UA was found in N. trachonitica (1.92 mg UA/g). The bioactivities of UA were evaluated with antioxidant and DNA protection activities, enzyme inhibitions, kinetics and interactions. The inhibition values (IC50) of α-amylase, α-glucosidase, urease, CA, tyrosinase, lipase, AChE, and BChE were determined between 5.08 and 181.96 µM. In contrast, Ki values of enzyme inhibition kinetics were observed between 0.04 and 0.20 mM. In addition, Ki values of these enzymes for enzyme-UA interactions were calculated as 0.38, 0.86, 0.45, 1.01, 0.23, 0.41, 0.01 and 2.24 µM, respectively. It is supported that UA can be widely used as a good antioxidant against oxidative damage, an effective DNA protector against genetic diseases, and a suitable inhibitor for metabolizing enzymes.Communicated by Ramaswamy H. Sarma.

Protective effect of alpha-lipoic acid, aerobic or resistance exercise from colitis in second hand smoke exposed young rats.

The role of second hand smoke (SHS) exposure on ulcerative colitis is not known. Our aim was to examine the effects of α-lipoic acid (ALA), chronic aerobic (AE) or resistance exercise (RE) on SHS exposed rats with colitis. Sprague-Dawley male rats (150-200 g, n=54) were selected for colitis induction. Among the colitis groups, one group was exposed to SHS (6 d/wk, 4 cigarettes/d) and the other was not. The SHS group was divided into subgroups as follows: sedentary; AE (swimming; 3 d/wk); and RE (climbing with weight; 3 d/wk). After 5 weeks, colitis was induced by intrarectal acetic acid. All groups had subgroups that were given subcutaneously ALA (50 mg/kg per day) or vehicle for 3 days. Following decapitation, colon tissues were sampled to examine malondialdehyde (MDA) and glutathione (GSH) levels, myeloperoxidase (MPO) activity, luminol and lucigenin chemiluminenscence, macroscopic scoring and histologic examination. ANOVA and Student's t-test were used for statistical analysis. The increased macroscopic and microscopic scores, MPO, MDA, luminol and lucigenin measurements in colitis and SHS-colitis groups were decreased via ALA (P<.05-.001). AE declined macroscopic and microscopic scores, MDA, lucigenin compared to colitis and SHS-colitis groups (P<.01-.001). RE reduced microscopic score, MPO, MDA, luminol, lucigenin (P<.05-.001) that were increased with colitis. Decreased GSH levels (P<.01) in the SHS-colitis group approached to control levels when given ALA. According to our results SHS and colitis induction increased inflammatory damage. SHS did not worsen it more than colitis. Our results suggest that ALA, AE or RE might be protective for SHS exposed ulcerative colitis conditions.