Characterization of Phenolic and Flavonoid Constituents from Citrus sinensis Peels with Antimicrobial Potentials: An Integrated In Vitro and In Silico Approach

Abstract

Introduction: Oranges (Citrus sinensis) are among the most widely cultivated citrus fruits, valued for their high vitamin C content and diverse bioactive compounds. While the edible portion is extensively used, the peel is often discarded as waste despite being a rich source of phytochemicals with potential therapeutic properties. This study focuses on the isolation and characterization of phenolics and flavonoids from orange peels and the evaluation of their antimicrobial potential through molecular docking and in vitro assays.

Materials and Methods :Methanolic extraction of orange peel was conducted using reflux condensation to obtain the methanolic extract of orange peel (ME-OP). Thin Layer Chromatography (TLC) and UV-visible spectroscopy were used for preliminary identification of phenolics and flavonoids. Structural elucidation was performed using Liquid Chromatography–Mass Spectrometry (LC-MS). Molecular docking studies were carried out to evaluate the interaction of isolated compounds with bacterial DNA gyrase (PDB ID: 3G75). Phytochemical screening and quantitative estimation of total phenolic and flavonoid content were also conducted, along with antimicrobial assays against selected Gram-positive and Gram-negative bacteria and fungi.

Results and Discussion : Phytochemical screening of ME-OP revealed the presence of carbohydrates, phytosterols, oils, proteins, amino acids, flavonoids, and abundant phenolics. TLC analysis showed flavonoids with Rf values of 0.17–0.23 and phenolics in the range of 0.33–0.63. Quantitative analysis indicated total phenolic content of 1.25 mg GAE/g and flavonoid content of 1.51 mg QE/g. LC-MS identified major phenolic acids such as p-coumaric, protocatechuic, caffeic, and ferulic acid, along with flavonoids including hesperidin and naringenin-7-O-glucoside. Molecular docking demonstrated that hesperidin and naringenin-7-O-glucoside had stronger binding affinities to bacterial DNA gyrase than the standard antibiotic ciprofloxacin. In vitro antimicrobial assays showed significant inhibition of Gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis), moderate activity against Gram-negative bacteria, and minimal antifungal effects.

Conclusions:This study highlights orange peel as a valuable source of bioactive phenolics and flavonoids with promising antimicrobial activity. The strong binding affinity of key flavonoids to bacterial targets and their effective inhibition of Gram-positive bacteria indicates the potential of orange peel extract as a natural antimicrobial agent. These findings support the therapeutic potential of citrus peel waste and encourage its utilization in the development of plant-based pharmaceuticals or antimicrobial formulations..