Anthocyanin-rich Berberis asiatica fruit extract: LC-MS based chemical profiling and evaluation of antimicrobial and antioxidant properties, and molecular docking study

Abstract

Aim: This study aimed to investigate the fruit ofBerberisasiatica as a potential source of bioactive anthocyanins and to evaluate their antioxidant and antimicrobial properties with insights into molecular docking studies. Methods: Crude extracts were prepared using solvents of varying polarity and characterized by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and Fourier-transform infrared spectroscopy (FT-IR) analyses. The total anthocyanin content was quantified, and antioxidant activity was assessed using the DPPH radical scavenging assay and total antioxidant capacity. Antimicrobial activity was evaluated against selected bacterial and fungal strains. Additionally, in silico molecular docking studies were performed to examine ligand-target interactions. Results: LC-MS/MS analysis identified eight compounds, including cyanidin-3-O-glucoside, cyanidin-3,5-diglucoside, malvidin-3-O-arabinoside, pelargonidin-3-O-glucoside, peonidin-3-O-glucoside, petunidin-3-O-glucoside, catechin, and epicatechin, indicating a pigment profile dominated by mono- and diglycosylated anthocyanins. The total anthocyanin content was 128.72 mg/g dry fruit, exceeding previously reported values for related species. Methanolic (80% v/v) and hydroalcoholic (50% v/v) extracts showed strong antioxidant activity (DPPH IC₅₀=10.13 and 12.56 µg/mL, respectively), whereas nonpolar fractions were less active. At 200 µg/mL, these extracts exhibited significant antimicrobial activity, with inhibition zones up to 42 mm againstEscherichiacoli and 41 mm againstMicrococcusluteus, along with antifungal effects againstAspergillusniger andCandidaalbicans. Docking studies revealed favorable binding energies (–7.3 to –8.0 kcal/mol) for key compounds against selected microbial and enzymatic targets. Conclusions: The findings demonstrate thatBerberisasiatica fruit is a rich source of anthocyanin-based pigments with potent antioxidant and antimicrobial activities. These results highlight its potential as a sustainable source of multifunctional bioactive compounds for nutraceutical and therapeutic applications.

Keywords

natural products / bioactive compounds / polar compounds / biological activity / anthocyanins

References

[1] Hoareau L, DaSilva EJ. Medicinal plants: a re—emerging health aid. Electron J Biotechnol. 1999;2:56-70.

[2] Subhose V, Srinivas P, Narayana A. Basic principles of pharmaceutical science in Ayurvĕda. Bull Indian Inst Hist Med Hyderabad. 1953;8:35-83.

[3] Thas JJ. Siddha medicine——background and principles and the application for skin diseases. Clin Dermatol. 2008;26:62-78.

[4] Ivan IM, Olaru OT, Popovici V, Chițescu CL, Popescu L, Luță EA, et al. Antioxidant and Cytotoxic Properties ofBerberisvulgaris (L.) Stem Bark Dry Extract . Molecules. 2024;29:2053.

[5] Salehi B, Selamoglu Z, Sener B, Kilic M, Kumar Jugran A, de Tommasi N, et al. Berberis Plants—Drifting from Farm to Food Applications, Phytotherapy, and Phytopharmacology . Foods. 2019;8:522.

[6] Dobhal P, Agnihotri S, Ashfaqullah S, Tamta S. A Review Study on Phytochemical and Pharmacological Aspects of Berberis asiatica Roxb. ex DC. Res Rev: J Pharm Sci. 2020;11:8-15.

[7] Castañeda—Ovando A, Pacheco—Hernández ML, Páez—Hernández ME, Rodríguez JA, Galán—Vidal CA. Chemical studies of anthocyanins: A review. Food Chem. 2009;113:859-71.

[8] He J, Giusti MM. Anthocyanins: natural colorants with health—promoting properties. Annu Rev Food Sci Technol. 2010;1:163-87.

[9] Gaur RD. Flora of the District Garhwal, North West Himalaya (With Ethnobotanical Notes). Taxon. 2000;49:840.

[10] Alade PI, Irobi ON. Antimicrobial activities of crude leaf extracts of Acalypha wilkesiana. J Ethnopharmacol. 1993;39:171—4.

[11] Mathur A, Bhat R, Prasad G, Dua VK, Verma SK, Agarwal PK. Antimicrobial activity of plants traditionally used as medicines against some pathogens. Rasayan J Chem. 2010;3:615—20.

[12] Aguila MC, McCann SM. Methods for the study of somatostatin. Methods Enzymol. 1989;168:716-29.

[13] Fargare T. In vitro antioxidant activity of plant extracts by DPPH assay. Eur Poly J. 1995;31:489-97.

[14] Pérez C, Anesini C. In vitro antibacterial activity of Argentine folk medicinal plants against Salmonella typhi. J Ethnopharmacol. 1994;44:41-6.

[15] Singleton VL, Rossi JA. Colorimetry of Total Phenolics with Phosphomolybdic—Phosphotungstic Acid Reagents. Am J Enol Vitic. 1965;16:144-58.

[16] Prieto P, Pineda M, Aguilar M. Spectrophotometric quantitation of antioxidant capacity through the formation of a phosphomolybdenum complex: specific application to the determination of vitamin E. Anal Biochem. 1999;269:337—41.

[17] Mathur A, Verma SK, Gupta V, Singh SK, Singh S, Mathur D, et al. Comparative studies on different varieties of apple (Pyrusmalus L.) of Kashmir (J&K) on the basis of PPO activity, Total Phenolic Content (TPC) andinvitro antioxidant activity . Pharma Sci Monit. 2011;2:S1-6.

[18] Trott O, Olson AJ. AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem. 2010;31:455-61.

[19] Pereira JH, Goh EB, Keasling JD, Beller HR, Adams PD. Structure of FabH and factors affecting the distribution of branched fatty acids in Micrococcus luteus. Acta Crystallogr D Biol Crystallogr. 2012;68:1320—8.

[20] Lafitte D, Lamour V, Tsvetkov PO, Makarov AA, Klich M, Deprez P, et al. DNA gyrase interaction with coumarin—based inhibitors: the role of the hydroxybenzoate isopentenyl moiety and the 5’—methyl group of the noviose. Biochemistry. 2002;41:7217—23.

[21] Bonin M, Hameleers L, Hembach L, Roret T, Cord—Landwehr S, Michel G, et al. In silico and in vitro analysis of an Aspergillus niger chitin deacetylase to decipher its subsite sugar preferences. J Biol Chem. 2021;297:101129.

[22] Cao H, Pauff JM, Hille R. Substrate orientation and catalytic specificity in the action of xanthine oxidase: the sequential hydroxylation of hypoxanthine to uric acid. J Biol Chem. 2010;285:28044—53.

[23] DeLano W. The PyMOL Molecular Graphics System. Version 0.99rc6. San Carlos: Schrödinger, LLC, De Lano Scientific; 2022.

[24] Pinzi L, Rastelli G. Molecular Docking: Shifting Paradigms in Drug Discovery. Int J Mol Sci. 2019;20:4331.

[25] Jara YS, Mekiso TT, Washe AP. Highly efficient catalytic degradation of organic dyes using iron nanoparticles synthesized with Vernonia Amygdalina leaf extract. Sci Rep. 2024;14:6997.

[26] Dara A, Feizy J, Naji—Tabasi S, Fooladi E, Rafe A. Intensified extraction of anthocyanins fromBerberisvulgaris L. by pulsed electric field, vacuum—cold plasma, and enzymatic pretreatments: modeling and optimization . Chem Biol Technol Agric. 2023;10:93.

[27] Pradhan PC, Saha S. Anthocyanin profiling of Berberis lycium Royle berry and its bioactivity evaluation for its nutraceutical potential. J Food Sci Technol. 2016;53:1205-13.

[28] Olivas—Aguirre FJ, Rodrigo—García J, Martínez—Ruiz ND, Cárdenas—Robles AI, Mendoza—Díaz SO, Álvarez—Parrilla E, et al. Cyanidin—3—O—glucoside: Physical—Chemistry, Foodomics and Health Effects. Molecules. 2016;21:1264.

[29] Prayoga DK, Aulifa DL, Budiman A, Levita J, Jiranusornkul S. Cyanidin and Cyanidin—3—Glucoside Alleviate Peptic Ulcer Disease: Insights from in vitro, and in vivo Studies. Drug Des Devel Ther. 2025;19:841-56.

[30] Płatosz N, Bączek N, Topolska J, Szawara—Nowak D, Wiczkowski W. The Blood—Cerebrospinal Fluid Barrier Features Different Permeability to Cyanidin—3—galactoside and Cyanidin—3—diglucoside—5—glucoside and Their Metabolites Circulating in Blood. J Agric Food Chem. 2022;70:12852-64.

[31] Yang S, Yang P, Wang W, Wu J, Liao X, Xu Z. A normalization strategy for cyanidin—3—diglucoside—5—glucoside recovery from red cabbage byproducts. J Cleaner Prod. 2025;494:144965.

[32] Xu Y, Ke H, Li Y, Xie L, Su H, Xie J, et al. Malvidin—3—O—Glucoside from Blueberry Ameliorates Nonalcoholic Fatty Liver Disease by Regulating Transcription Factor EB—Mediated Lysosomal Function and Activating the Nrf2/ARE Signaling Pathway . J Agric Food Chem. 2021;69:4663-73.

[33] Merecz—Sadowska A, Sitarek P, Kowalczyk T, Zajdel K, Jęcek M, Nowak P, et al. Food Anthocyanins: Malvidin and Its Glycosides as Promising Antioxidant and Anti—Inflammatory Agents with Potential Health Benefits. Nutrients. 2023;15:3016.

[34] Ren Z, Raut NA, Lawal TO, Patel SR, Lee SM, Mahady GB. Peonidin—3—O—glucoside and cyanidin increase osteoblast differentiation and reduce RANKL—induced bone resorption in transgenic medaka. Phytother Res. 2021;35:6255-69.

[35] Sari DRT, Cairns JRK, Safitri A, Fatchiyah F. Virtual Prediction of the Delphinidin—3—O—glucoside and Peonidin—3—O—glucoside as Anti—inflammatory of TNF—α Signaling. Acta Inform Med. 2019;27:152—7.

[36] Karim N, Shishir MRI, Li Y, Zineb OY, Mo J, Tangpong J, et al. Pelargonidin—3—O—Glucoside Encapsulated Pectin—Chitosan—Nanoliposomes Recovers Palmitic Acid—Induced Hepatocytes Injury . Antioxidants (Basel). 2022;11:623.

[37] Semmarath W, Mapoung S, Umsumarng S, Arjsri P, Srisawad K, Thippraphan P, et al. Cyanidin—3—O—glucoside and Peonidin—3—O—glucoside—Rich Fraction of Black Rice Germ and Bran Suppresses Inflammatory Responses from SARS—CoV—2 Spike Glycoprotein S1—Induction In Vitro in A549 Lung Cells and THP—1 Macrophages via Inhibition of the NLRP3 Inflammasome Pathway. Nutrients. 2022;14:2738.

[38] Wang X, Wang W, Luo S, Wang N, Wang L, Zhang N, et al. Evaluation of ohmic heating modified soybean protein isolate structure and antioxidant film under different catechin concentrations. LWT. 2023;186:115224.

[39] German IJS, Pomini KT, Andreo JC, Shindo JVTC, Castro MVM, Detregiachi CRP, et al. New Trends to Treat Muscular Atrophy: A Systematic Review of Epicatechin. Nutrients. 2024;16:326.

[40] Huang W, Zhang S, Qin G, Wenquan G, Wu J. Isolation and determination of major anthocyanin pigments in the pericarp ofP.communis L. cv. ‘Red Du Comices’ and their association with antioxidant activity . African J Agric Res. 2012;7:3772-80.

[41] Wu X, Prior RL. Identification and characterization of anthocyanins by high—performance liquid chromatography—electrospray ionization—tandem mass spectrometry in common foods in the United States: vegetables, nuts, and grains. J Agric Food Chem. 2005;53:3101—13.

[42] Wu X, Prior RL. Systematic identification and characterization of anthocyanins by HPLC—ESI—MS/MS in common foods in the United States: fruits and berries. J Agric Food Chem. 2005;53:2589-99.

[43] Mazzca P, Ferranti P, Picariello G, Chianese L, Addeo F. Mass spectrometry in the study of anthocyanins and their derivatives: differentiation of Vitis vinifera and hybrid grapes by liquid chromatography/electrospray ionization mass spectrometry and tandem mass spectrometry. J Mass Spectrom. 2005;40:83-90.

[44] Liu GL, Guo HH, Sun YM. Optimization of the extraction of anthocyanins from the fruit skin of Rhodomyrtus tomentosa (Ait.) Hassk and identification of anthocyanins in the extract using High—Performance Liquid Chromatography—Electrospray Ionization—Mass Spectrometry (HPLC—ESI—MS). Int J Mol Sci. 2012;13:6292—302.

[45] Hutabarat RP, Xiao YD, Wu H, Wang J, Li DJ, Huang WY. Identification of Anthocyanins and Optimization of Their Extraction from Rabbiteye Blueberry Fruits in Nanjing. J Food Qual.2019;2019:1-10.

[46] Bhandari DK, Nath G, Ray AB, Tewari PV. Antimicrobial activity of crude extracts from berberis asiatica stem bark. Pharm Biol. 2000;38:254—7.

[47] Bhattarai K, Dhakal D, Pandey I, Subba B, Sharma KR. Phytochemistry, Biological Activities, and Chemical Profiling of Berberis asiatica. BIBECHANA. 2023;20:236-47.

[48] Singh M, Srivastava S, Rawat A. Antimicrobial activities of IndianBerberis species . Fitoterapia. 2007;78:574-6.

[49] Stein—Chisholm R, Beaulieu J, Grimm C, Lloyd S. LC—MS/MS and UPLC—UV Evaluation of Anthocyanins and Anthocyanidins during Rabbiteye Blueberry Juice Processing. Beverages. 2017;3:56.

[50] Malik Z, Jain K, Ravindran K, Sathiyaraj G. In vitro antimicrobial activity and preliminary phytochemical analysis of Berberis aristata. J Ethnobiol Ethnomed. 2017;4:1-6.

[51] Anubhuti P, Rahul S, Chandra Kant K. Comparative study on the antimicrobial activity of Berberis aristata from different regions and Berberine in vitro. Int J Life Sci Pharma Res. 2011;1:L17-20.

[52] Sun J, Li Q, Li J, Liu J, Xu F. Nutritional composition and antioxidant properties of the fruit of Berberis heteropoda Schrenk. PLoS One. 2022;17:e0262622.

[53] Ahmed M, Bose I, Goksen G, Roy S. Himalayan Sources of Anthocyanins and Its Multifunctional Applications: A Review. Foods. 2023;12:2203.

[54] Belwal T, Pandey A, Bhatt ID, Rawal RS, Luo Z. Trends of polyphenolics and anthocyanins accumulation along ripening stages of wild edible fruits of Indian Himalayan region. Sci Rep. 2019;9:5894.