Objective: This study aimed to evaluate the arginase inhibitory activity of Caesalpinia tortuosa Roxb. stem bark extracts.Methods: C. tortuosa Roxb. stem bark extracts were obtained through reflux extraction using n-hexane, ethyl acetate, and methanol and their inhibitory activity against arginase was measured using a microplate reader at 430 nm. Active extracts were subjected to phytochemical analysis and based on the qualitative phytochemical analysis, quantitative data regarding flavonoid and phenolic contents were obtained. The total flavonoid content of active extracts was determined using AlCl 3 colorimetric method, and the phenolic content was determined using Folin-Ciocalteu method. Results:Ethyl acetate and methanol extracts of C. tortuosa Roxb. inhibited activity of arginase with IC 50 values of 33.81 and 11.58 μg/mL, respectively, nor-NOHA acetate as standard drug inhibited arginase with IC 50 values of I3.77 μg/mL. Both active extracts contained saponins, tannins, and flavonoids. Ethyl acetate and methanol extracts showed a total flavonoid content of 7.41 mgQE/g and 5.05 mgQE/g and total phenolic content of 27.55 mgGE/g and 17.16 mgGE/g, respectively. Methanol extracts had a higher inhibitory activity than ethyl acetate extracts despite having flavonoid and phenolic content, thereby suggesting no correlation between arginase inhibitory activity and flavonoid or phenolic content. Conclusion:Ethyl acetate and methanolic extracts of C. tortuosa Roxb. stem barks containing flavonoids, tannins, and saponins displayed arginase inhibitory activity, and no correlation was observed between arginase inhibitory activity and flavonoid and phenolic content.Substrate optimization was performed using concentrations of 130, 570, 650, and 820 mM as suggested by the protocol. Substrate concentrations were tested using 1 U/mL of arginase enzyme. The procedure was performed in triplicate using 10 µL of bidistillation water, 15 µl of enzyme solution, and 20 µl of substrate solution followed by incubation for 30 min at 37°C. After incubation, 100 µl of urea was directly added, followed by incubation at room temperature (25°C).
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