Pre-Clinical In-Vivo and In-Vitro Methods For Evaluation of Anti-Alzheimer’s Drugs
Atanu Bhattacharjee, Akula Ramakrishna, Magisetty Obulesu in Phytomedicine and Alzheimer’s Disease, 2020
The enzyme activity is calculated using the following equation: whereA/min= Change in absorbance per minε = 1.361 ×104 M–1 cm–1b = path length (1-cm)Vt = total volume (3.1 ml)Vs = sample volume (0.4 ml)
General concepts for applied exercise physiology
Nick Draper, Helen Marshall in Exercise Physiology, 2014
The velocity or rate at which an enzyme catalyses a reaction is dependent on a number of factors including temperature (as already mentioned), substrate concentration, enzyme concentration, cellular pH and ionic strength of the aqueous medium. Figure 8.12 provides an illustration of the effect of increasing temperature, substrate concentration and pH on enzyme activity. Enzymes cease to catalyse reactions at temperatures close to 0, but for each 10°C increase in temperature up to around 45–50°C, enzyme activity doubles. This is known as the Q10 quotient. The increase in enzymatic activity, which increases the reaction rate, occurs because the increase in temperature leads to an increase in the number of collisions between the substrate and the enzyme. Beyond 45–50°C, however, enzymes denature, which changes the shape of the protein and sharply decreases catalytic activity. This means that enzymes are ideally suited to catalysing biochemical reactions. This is one of the reasons why warm-ups are beneficial for sports performance. Enzyme activity and therefore ATP synthesis are enhanced by the increased muscle temperatures associated with a thorough warm-up.
Alcohols and Aldehydes
Frank A. Barile in Barile’s Clinical Toxicology, 2019
Ethanol elimination depends on the overall enzyme rate for ADH activity for the metabolism of ethanol. Figure 24.2 illustrates the plot of ADH activity versus ethanol concentration (per unit time, usually hours). At or below the maximum enzyme rate of about 23 mg/dl (Vmax) for ADH, ethanol metabolism (and elimination) follows first-order kinetics; that is, the amount of ethanol metabolized is directly proportional to the concentration. Above the Vmax for the enzyme, ethanol metabolism follows zero-order kinetics, whereby a further increase in ethanol concentration is not followed by a proportionate increase in enzyme activity. Thus, ethanol metabolism remains constant above Vmax even as the concentration increases. Based on these rates, the equilibrium constant (or average enzyme rate constant, Km) is calculated as
Investigation of the effects of cephalosporin antibiotics on glutathione S-transferase activity in different tissues of rats in vivo conditions in order to drug development research
Published in Drug and Chemical Toxicology, 2020
Fikret Türkan, Zübeyir Huyut, Parham Taslimi, Mehmet Tahir Huyut, İlhami Gülçin
In recent years, inhibition studies of many antibiotics have been performed extensively. These studies were conducted on various enzymes, including paraoxonase (Demir and Beydemir 2015, Türkeş et al.2015). glucose-6-phosphate dehydrogenase (Ozmen et al.2005), 6-phosphogluconate dehydrogenase (Akyüz et al.2004), glutathione reductase (Erat et al.2005), and glutathione S-transferase (Comakli et al.2011). In vivo results from the enzyme activity studies are crucial to recognize the physiological role of the enzyme. Particularly, drug-enzyme or any chemical compound-enzyme interaction studies are important to understand the toxicological mechanisms. In present study, we evaluated cefazolin, cefuroxime, and cefoperazone in vivo inhibition effect on GST enzyme activity. In cefazolin and cefoperazone groups, GST enzyme activities were increased in liver and renal tissues during the first 5 h period, and then began to drop in the next time period. In addition, they were increased in cefuroxime group during firs 3 h period, while began to drop in the next time periods. The situation was showed that the cefazoline and cefaperazon were metabolized after 5th hour and completed their half-life, while cefuroxime were metabolized after 3th hour on GST activity. In addition, after 7 h, it was observed that cefazolin had no adverse effect on GSH enzyme activity in all working tissues, but cefuroxime and cefaperazon caused the decreasing in GST activities compared to control groups after 5th hour period.
Inhibition of Echis ocellatus venom metalloprotease by flavonoid-rich ethyl acetate sub-fraction of Moringa oleifera (Lam.) leaves: in vitro and in silico approaches
Published in Toxin Reviews, 2022
Akindele Oluwatosin Adeyi, Kaosarat Keji Mustapha, Babafemi Siji Ajisebiola, Olubisi Esther Adeyi, Damilohun Samuel Metibemu, Raphael Emuebie Okonji
The effects of temperature on enzyme activity were carried out between 30 °C and 100 °C. The assay mixture was first incubated at the selected temperature for 10 min before the reaction was initiated by the addition of 50 µl enzyme that had been incubated at the same temperature. For heat stability of the enzyme, an aliquot of the enzyme was taken and incubated at temperatures of 40, 50, 60, 70, and 80 °C for a period of 1 h and at intervals of 10 min, the enzyme was taken and assayed for residual activity. The effect of pH on metalloproteinase activity was assayed at different pH values. The pH values were varied by using buffers of different pH, which includes 0.01 M citrate buffer (pH 3.0–5.0), 0.01 M Phosphate buffer (pH 6.0–7.0), 1 mM Tris buffer (pH 8.0) and borate buffer (pH 9.0–10.0). Reaction mixture each contained 1 mM of a different buffer, 0.33 M of arginine, and 0.05 ml of the enzyme (Lee et al.2014).
Specific and combined effects of dietary ethanol and arginine on Drosophila melanogaster
Published in Drug and Chemical Toxicology, 2023
Maria M. Bayliak, Oleh I. Demianchuk, Dmytro V. Gospodaryov, Vitalii A. Balatskyi, Volodymyr I. Lushchak
Glucose-6-phosphate dehydrogenase activity was measured by monitoring NADP+ reduction at 340 nm in a reaction mixture containing 50 mM KPi, pH 7.5, 5 mM MgCl2, 2 mM glucose-6-phosphate, 0.2 mM NADP+, and 40 µl of supernatant in a final volume of 1 ml (Lushchak et al. 2011). The extinction coefficient for NADPH of 6.22 mM−1 cm−1 was used for calculation of G6PDH activity. Glutathione-S-transferase activity was measured by monitoring formation of an adduct of GSH and 1-chloro-2,4-dinitrobenzene (CDNB) at 340 nm. Reaction mixture of GST measurement contained 50 mM Kpi (pH 7.0), 0.5 mM EDTA, 5 mM GSH, 1 mM CDNB, and 3–5 μl of supernatant in a final volume of 1 ml (Gospodaryov et al. 2020). The extinction coefficient for an adduct of GSH and CDNB of 9.6 mМ−1 сm−1 was used for calculation of GST activity. One unit of enzyme activity was defined as the amount of the enzyme generating 1 μmol of product per minute; activities are expressed as international units or milliunits per milligram of soluble protein. Protein concentrations were determined by the Bradford assay (Bradford 1976).
Related Knowledge Centers
- Enzyme
- Enzyme Inhibitor
- Enzyme Kinetics
- Gelatin
- Turnover Number
- Substrate
- Assay
- Flash Photolysis
- Stopped-Flow
- Temperature Jump