Molecular docking and inhibition profiles of some antibiotics on lactoperoxidase enzyme purified from bovine milk.

Antibiotics are generally used for human and veterinary applications to preserve and to control microbial diseases. Milk has a biologically significant enzyme known as lactoperoxidase (LPO) that is a member of peroxidase family. In metabolism, LPO has ability to catalyze the transformation of thiocyanate (SCN-) to hypothiocyanite (OSCN-) that is an antibacterial agent and the reaction occurs with hydrogen peroxide. In this work, LPO inhibition effects of some antibiotics including cefazolin, oxytetracycline, flunixin meglumine, cefuroxime, tylosin, vancomycin, chloramphenicol and lincomycin were tested. Among the antibiotics cefazolin was indicated the strongest inhibitory efficacy. The half maximal inhibitory concentration (IC50) and the inhibition constant (Ki) values of cefazolin were found as 8.19 and 34.66 µM, respectively. It was shown competitive inhibition. 5-Methyl-1,3,4-thiadiazol-2-yl moiety activity plays a key role in the inhibition mechanism of cefazolin.Communicated by Ramaswamy H. Sarma.

Some indazoles as alternative inhibitors for potato polyphenol oxidase.

Fresh-cut vegetables and fruits have gained attention among consumers because of their fresh appearance, lack of pollution, nutrition, and convenience. However, in fresh-cut foods, enzymatic browning is the main problem. Polyphenol oxidase (PPO) is a vital enzyme involved in the process of enzymatic browning. In this study, PPO was purified from potato using Sepharose 4B-l-tyrosine-p-aminobenzoic acid affinity chromatography and the effect of some indazoles on the enzyme was determined. The enzyme was purified with a specific activity of 52,857.14 EU/mg protein and 21.26-purification fold. Indazoles exhibited inhibitor properties for PPO with IC50 values in the range of 0.11-1.12 mM and Ki values in the range of 0.15 ± 0.04-3.55 ± 0.88 mM. Among these compounds, 7-chloro-1H-indazole was shown as the most potent PPO inhibitor (Ki : 0.15 ± 0.04 mM). Determination of the enzyme's inhibition kinetics will simplify the testing of candidate PPO inhibitors.

Purification of Polyphenol Oxidase from Potato and Investigation of the Inhibitory Effects of Phenolic Acids on Enzyme Activity.

BACKGROUND: Polyphenol Oxidase (PPO) belongs to the oxidoreductase enzyme family. METHODS: Here, PPO was purified from potato using Sepharose 4B-L-tyrosine-p-aminobenzoic acid affinity chromatography. It determined the interactions between some phenolic acids and the enzyme. RESULTS: The enzyme was obtained with a specific activity of 15333.33 EU/mg protein and 7.87- fold purification. It was found that phenolic acids exhibited inhibitory properties for PPO. The IC50 values of the phenolic acids were found in the range of 0.36-2.12 mM, and their Ki values were found in the range of 0.28± 0.07-1.72±0.32 mM. It was determined that all studied compounds displayed a competitive inhibition effect. Among these compounds, 3-hydroxybenzoic acid was found to be the most effective PPO inhibitor (Ki: 0.28±0.07 mM). CONCLUSION: Investigating the inhibition kinetics of the enzyme will simplify the testing of PPO inhibitor candidates.

Investigation of the effects of some sulfonamides on acetylcholinesterase and carbonic anhydrase enzymes.

Human carbonic anhydrase I and II isoenzymes (hCA I and II) and acetylcholinesterase (AChE) are important metabolic enzymes that are closely associated with various physiological and pathological processes. In this study, we investigated the inhibition effects of some sulfonamides on hCA I, hCA II, and AChE enzymes. Both hCA isoenzymes were purified by Sepharose-4B-L-Tyrosine-5-amino-2-methylbenzenesulfonamide affinity column chromatography with 1393.44 and 1223.09-folds, respectively. Also, some inhibition parameters including IC50 and Ki values were determined. Sulfonamide compounds showed IC 50 values of in the range of 55.14 to 562.62 nM against hCA I, 55.99 to 261.96 nM against hCA II, and 98.65 to 283.31 nM against AChE. Ki values were in the range of 23.40 ± 9.10 to 365.35 ± 24.42 nM against hCA I, 45.87 ± 5.04 to 230.08 ± 92.23 nM against hCA II, and 16.00 ± 45.53 to 157.00 ± 4.02 nM against AChE. As a result, sulfonamides had potent inhibition effects on these enzymes. Therefore, we believe that these results may contribute to the development of new drugs particularly in the treatment of some disorders.

Improved chromatographic method for purification of lactoperoxidase from different milk sources.

Our previous studies showed that sulfanilamide is a new competitive inhibitor of and can be used in the purification of lactoperoxidase (LPO, EC1.11.1.7) from milk. However, this method has some disadvantages like a lower purification factor. The aim of the present study is to improve the purification process of milk LPO from different sources. For this purpose, 16 commercial sulfanilamide derivatives were selected for inhibition studies to determine the best inhibitor of bovine LPO by calculating kinetic parameters. A cyanogen bromide-activated Sepharose 4B affinity matrix was synthesized by coupling with each competitive inhibitor. Among the inhibitors, 5-amino-2-methylbenzenesulfonamide and 2-chloro-4-sulfamoylaniline were used as ligands for the purification of LPO from bovine, buffalo, cow, and goat milks with 1059.37, 509.09, 232.55, and 161.90, and 453.12-, 151.86-, 869.00-, and 447.57-fold, respectively. Our results show that 5-amino-2-methylbenzenesulfonamide, 2-chloro-4-sulfamoylaniline, and 5-amino-1-naphthalenesulfonamide are the best inhibitors for one-step purification of the enzyme.

The relationship among pressure ulcers, oxygenation, and perfusion in mechanically ventilated patients in an intensive care unit.

PURPOSE: We examined the relationships among oxygenation, tissue perfusion, and other comorbid conditions not incorporated into the Norton Scale, and pressure ulcer (PU) development in subjects receiving mechanical ventilation. DESIGN: Descriptive, observational study. SETTING AND SUBJECTS: The setting was our university hospital's surgical/emergency intensive care unit in Istanbul province, Turkey. The sample comprised 30 patients who were older than 18 years, did not have a PU on admission, and had been mechanically ventilated for more than 24 hours when data collection began. METHODS: Skin integrity and a PU risk, using the Norton Scale, were administered twice daily. In addition, serum blood testing, vital signs, and data regarding ventilation and oxygenation status were obtained from the patient's electronic medical records. RESULTS: Slightly less than half of subjects were women (n = 14, 46.7%). Their mean age was 54.36 years (SD = 20.68). Pressure ulcers developed in 5 patients (16.7%); all PUs were located on the heel. All ulcers were initially observed as stage I lesions; 1 progressed to a stage II ulcer and 1 progressed to a stage III ulcer. Patients who developed pressure ulcers have higher serum glucose levels (z = -2.198; P = .028), higher serum pH levels (z = -2.031; P = .028), and lower diastolic blood pressures (z = 0.055; P = .057) than those who remained ulcer free. CONCLUSION: Our results demonstrate that mechanically ventilated patients who develop PUs were more likely to have significantly higher blood glucose levels, significantly lower diastolic blood pressure values, and significantly higher serum pH values than were patients who remained free of PUs. Nurses who care for mechanically ventilated patients should recognize these factors and initiate preventive interventions as indicated.