Chemical Profiling of Acacia auriculiformis (A. Cunn.) ex Benth. Leaves Extract and its Impact on Gene Expression and Apoptotic DNA Damage in CCl4-Induced Hepatotoxicity in Male Rats


  • Eman M. M. El-Taher Department of Pharmacognosy, Faculty of Pharmacy, Egyptian Russian University, Badr City, 63514 Cairo, Egypt
  • Moshera M. El-Sherei Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, 11562 Cairo, Egypt
  • Riham S. El Dine Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, 11562 Cairo, Egypt
  • Dina M. Y. El Naggar Department of Pharmacognosy, Faculty of Pharmacy, Egyptian Russian University, Badr City, 63514 Cairo, Egypt
  • Wagdy K. B. Khalil Department of Cell Biology, National Research Centre (NRC), 12622 Dokki, Giza, Egypt
  • Mariam G. Eshak Department of Cell Biology, National Research Centre (NRC), 12622 Dokki, Giza, Egypt
  • El-Sayed S. Abdel-Hameed Laboratory of Medicinal Chemistry, Theodor Bilharz Research Institute, Giza, Egypt
  • Mona E. S. Kassem Department of Phytochemistry and Plant Systematics, National Research Centre (NRC), 12622 Dokki, Giza, Egypt


CYP450, Hsp70, Genotoxicity, LC-ESI-MS, Acacia


Acacia auriculiformis (A. Cunn.) ex Benth. is an ornamental evergreen tree with diverse medicinal importance. It was subjected to different chromatographic techniques. Eighteen known phenolics were identified from the leaves using different spectroscopic methods. Among them, twelve compounds were isolated and characterized as: three phenolic acids, one flavone, four flavonols, three isoflavones, and one flavanone. From the LC-ESI-MS profiling technique additional six flavonoids were tentatively identified. HPLC-UV chromatographic quantitation for quercetin 3-O-glucoside, eriodicotyl 7-O-glucoside, genistein and biochanin A was determined as 3.475, 2.406, 6.658 and 5.997 mg/g defatted extract, respectively. The extract genoprotective effect was assessed in 80 adult male rats (10 rats/group) against CCl4-induced genotoxicity in liver tissues (1 mg/kg bw, twice per week), in three doses (35, 70 and 140 mg/kg
bw/d, four weeks), using real-time PCR, comet assay, DNA fragmentation and flow cytometry apoptosis assay. In treated rats, the plant extract significantly down-regulated the hepatic mRNA expression levels of cytochrome P450 (CYP 450) and heat shock protein 70 (Hsp 70) genes and attenuated DNA damage rates to 19.4, 16.2 and 13.8% for the three doses, respectively compared to that of the untreated (23.8%). It also reduced apoptotic DNA fragmentation and apoptotic rates by 26.2, 19.4 and 15.2%, respectively compared to CCl4 treated rats (37.1%). These findings indicated the potential effect of A. auriculiformis leaves to retain DNA structural integrity and its genomic expression against common industrial pollutant such as CCl4 in a dosedependent manner.


Seigler DS. Phytochemistry of Acacia—sensu lato. Biochem. Syst Ecol. 2003; 31(8):845-873.

Maslin B, Miller J, Seigler D. Overview of the generic status of Acacia (Leguminosae: Mimosoideae). Aust Syst Bot. 2003; 16(1):1-18.

Sathya A and Siddhuraju P. Role of phenolics as antioxidants, biomolecule protectors and as anti–diabetic factors–Evaluation on bark and empty pods of Acacia auriculiformis. Asian Pac J Trop Med. 2012; 5(10):757- 765.

Sharma N, Singh S, Singh SK. Review on Phytopharmacological Properties of Acacia auriculiformis A. Cunn. ex. Benth. Planta Activa 2016; 1:1-6.

Sharma N, Singh S, Singh SK. Pharmacognostical standardization and preliminary phytochemical investigations on Acacia auriculiformis A. Cunn. Ex. Benth stem bark. J Med Plant Res. 2017; 5(1):398-402.

Mandal P, Babu SS, Mandal N. Antimicrobial activity of saponins from Acacia auriculiformis. Fitoterapia 2005; 76(5):462-465.

Sathya A and Siddhuraju P. Protective effect of bark and empty pod extracts from Acacia auriculiformis against paracetamol intoxicated liver injury and alloxan induced type II diabetes. Food Chem Toxicol. 2013; 56:162-170.

Yang SF, Chang CW, Wei RJ, Shiue YL, Wang SN, Yeh YT. Involvement of DNA damage response pathways in hepatocellular carcinoma. BioMed Res Int. 2014; 2014:153867-153867.

Markham KR. Techniques of flavonoid identification. London: Academic press; 1982. 36 p.

Agrawal P and Bansal M. Flavonoids glycosides in “Carbon-13 NMR of Flavonoids. Amsterdam: Elsevier; 1989. 321 p.

Markham KR and Geiger H. 1H nuclear magnetic resonance spectroscopy of flavonoids and their glycosides in hexadeutero dimethyl sulphoxide. In: Harborne JB, editor. The flavonoids-advances in research since 1986. London: Chapman and Hall; 1994.

Ibrahim LF, Elkhateeb A, Marzouk MM, Hussein SR, Abdel-Hameed E-SS, Kassem ME. Flavonoid investigation, LC–ESIMS profile and cytotoxic activity of Raphanus raphanistrum L.(Brassicaceae). J Chem Pharm Res. 2016; 8(7):786-793.

Okokon JE, Jackson O, Opara KN, Emmanuel E. In vivo antimalarial activity of ethanolic leaf extract of Acacia Auriculiformis. Int J Drug Dev Res. 2010; 2:482-487.

Linjawi SA, Salem LM, Khalil WK. Jatropha curcas L. kernel prevents benzene induced clastogenicity, gene expression alteration and apoptosis in liver tissues of male rats. Indian J Exp Biol. 2017; 55:225-234.

El-Baz FK, Khalil WK, Booles HF, Aly HF, Ali GH. Dunaliella salina suppress oxidative stress, alterations in the expression of pro-apoptosis and inflammation related genes induced by STZ in diabetic rats. Int J Pharm Sci Rev Res. 2016; 38(02):219-226.

Kassem MES, Ibrahim LF, Hussein SR, El-Sharawy R, ElAnsari MA, Hassanane MM, Booles HF. Myricitrin and bioactive extract of Albizia amara leaves: DNA protection and modulation of fertility and antioxidant-related genes expression. Pharm Biol. 2016; 54(11):2404-2409.

Olive PL and Banáth JP. The comet assay: a method to measure DNA damage in individual cells. Nat Protoc. 2006; 1(1):23.

Lu T, Xu Y, Mericle MT, Mellgren RL. Participation of the conventional calpains in apoptosis. Bba-Mol Cell Res. 2002; 1590(1-3):16-26.

Schaffer S, Podstawa M, Visioli F, Bogani P, Müller WE, Eckert GP. Hydroxytyrosol-rich olive mill wastewater extract protects brain cells in vitro and ex vivo. J Agric Food Chem. 2007; 55(13):5043-5049.

Backler F and Wang F. Impact of intramolecular hydrogen bonding of gallic acid conformers on chemical shift through NMR spectroscopy. J Mol Graph Model. 2020; 95:107486.

Aung HT, Furukawa T, Nikai T, Niwa M, Takaya Y. Contribution of cinnamic acid analogues in rosmarinic acid to inhibition of snake venom induced hemorrhage. Bioorg Med Chem. 2011; 19(7):2392-2396.

Forino M, Tartaglione L, Dell’Aversano C, Ciminiello P. NMR-based identification of the phenolic profile of fruits of Lycium barbarum (goji berries). Isolation and structural determination of a novel N-feruloyl tyramine dimer as the most abundant antioxidant polyphenol of goji berries. Food chem. 2016; 194:1254-1259.

Kurkin VA, Morozova TV, Pravdivtseva OE, Kurkina AV, Daeva ED, Kadentsev VI. Constituents from Leaves of Crataegus sanguinea. Chem Nat Compd. 2019; 55(1):21- 24.

Safwat NA, Kashef MT, Aziz RK, Amer KF, Ramadan MA. Quercetin 3-O-glucoside recovered from the wild Egyptian Sahara plant, Euphorbia paralias L., inhibits glutamine synthetase and has antimycobacterial activity. Tuberculosis. 2018; 108:106-113.

Imperato F. Kaempferol 3-O-(acetylrutinoside), a New Flavonoid and Two New Fern Constituents, Quercetin 3-O- (acetylglucoside) And 3-O-(acetylrutinoside) From Dryopteris Villarii. Am Fern J. 2006; 96(03):93-96.

Pasayeva L, Üstün O, Demirpolat E, Karatoprak GS, Tugay O, Kosar M. Bioactivity-guided isolation of cytotoxic and antioxidant phytochemicals from four Cousinia species from Stenocephala bunge section. Pharmacogn Mag. 2019; 15(65):682-692.

Zhao S, Zhang L, Gao P, Shao Z. Isolation and characterisation of the isoflavones from sprouted chickpea seeds. Food Chem. 2009; 114(3):869-873.

Walters NA, de Villiers A, Joubert E, de Beer D. Improved HPLC method for rooibos phenolics targeting changes due to fermentation. J Food Compost Anal. 2017; 55:20-29.

Ioanna V and Ioannis P. Quality assessment and quantification of genistein in dietary supplements by highperformance liquid chromatography, quantitative nuclear magnetic resonance, and two-dimensional diffusion ordered spectroscopy 1H. Asian J Pharm Clin Res. 2020; 13(3):132- 142.

Sartorelli P, Carvalho CS, Reimão JQ, Ferreira MJP, Tempone AG. Antiparasitic activity of biochanin A, an isolated isoflavone from fruits of Cassia fistula (Leguminosae). Parasitol Res. 2009; 104(2):311-314.

Ferreres F, Gil-Izquierdo A, Vinholes J, Silva ST, Valentão P, Andrade PB. Bauhinia forficata Link authenticity using flavonoids profile: Relation with their biological properties. Food Chem. 2012; 134(2):894-904.

Santos-Zea L, Gutiérrez-Uribe JA, Serna-Saldivar SO. Comparative analyses of total phenols, antioxidant activity, and flavonol glycoside profile of cladode flours from different varieties of Opuntia spp. J Agric Food Chem. 2011; 59(13):7054-7061.

Farag MA, Sakna ST, El-fiky NM, Shabana MM, Wessjohann, LA. Phytochemical, antioxidant and antidiabetic evaluation of eight Bauhinia L. species from Egypt using UHPLC–PDA–qTOF-MS and chemometrics. Phytochem. 2015; 119:41-50.

Lu Y, Zhu S, He Y, Mo C, Wu C, Zhang R, Zheng X, Tang Q. Systematic characterization of flavonoids from Siraitia grosvenorii leaf extract using an integrated strategy of high‐ speed counter‐current chromatography combined with ultra high performance liquid chromatography and electrospray ionization quadrupole time‐of‐flight mass spectrometry. J Sep Sci. 2020; 43(5):852-864.

Diab KA, Fahmy MA, Hassan ZM, Hassan EM, Salama AB, Omara EA. Genotoxicity of carbon tetrachloride and the protective role of essential oil of Salvia officinalis L. in mice using chromosomal aberration, micronuclei formation, and comet assay. Environ Sci Pollut Res. 2018; 25(2):1621- 1636.

Nofal ZAE, El-Maghraby AF, Gad MR, Farghaly AA, Ahmed EM. Evaluation of the Protective Effect of Ethanolic Extract of Plantago major Plant Against CCl4 Induced Genotoxicity and Biochemical Disturbance in Rat. Egyptian J Vet Sci. 2016; 47(1):83-110.

He L, He T, Farrar S, Ji L, Liu T, Ma X. Antioxidants maintain cellular redox homeostasis by elimination of reactive oxygen species. Cell Physiol Biochem. 2017; 44(2):532-553.

Holden PR, James NH, Brooks AN, Roberts RA, Kimber I, Pennie WD. Identification of a possible association between carbon tetrachloride–induced hepatotoxicity and interleukin‐8 expression. J Biochem Mol Toxicol. 2000; 14(5):283-290.

Godard CA, Leaver MJ, Said MR, Dickerson RL, George S, Stegeman JJ. Identification of cytochrome P450 1B-like sequences in two teleost fish species (scup, Stenotomus chrysops and plaice, Pleuronectes platessa) and in a cetacean (striped dolphin, Stenella coeruleoalba). Mar

Environ Res. 2000; 50(1-5):7-10.

Kurtz AE, Reiner JL, West KL, Jensen BA. Perfluorinated alkyl acids in Hawaiian cetaceans and potential biomarkers of effect: peroxisome proliferator-activated receptor alpha and cytochrome P450 4A. Environ Sci 2019; 53(5):2830- 2839.

Jacob P, Hirt H, Bendahmane A. The heat‐shock protein/chaperone network and multiple stress resistance. Plant Biotechnol J. 2017; 15(4):405-414.

Zhu QY, Hackman RM, Ensunsa JL, Holt RR, Keen CL. Antioxidative activities of Oolong tea. J Agric Food Chem. 2002; 50(23):6929-6934.

Losada-Barreiro S and Bravo-Diaz C. Free radicals and polyphenols: The redox chemistry of neurodegenerativediseases. Eur J Med Chem. 2017; 133:379-402.

Pool-Zobel, Beatrice L., Herman Adlercreutz, Michael Glei, Ute M. Liegibel, Julie ittlingon, an Rowland, ristiina h l , erhard Rechkemmer. soflavonoids and lignans have different potentials to modulate oxidative genetic damage in human colon cells. Carcinogenesis. 2000; 21(6):


Jomova K, Lawson M, Drostinova L, Lauro P, Poprac P, Brezova V, Michalik M, Lukes V, Valko M. Protective role of quercetin against copper (II)-induced oxidative stress: A spectroscopic, theoretical and DNA damage study. Food Chem Toxicol. 2017; 110:340-350.

Kahkeshani N, Farzaei F, Fotouhi M, Alavi SS, Bahramsoltani R, Naseri R, Momtaz S, Abbasabadi Z, Rahimi R, Farzaei MH, Bishayee A. Pharmacologicaleffects of gallic acid in health and diseases: A mechanistic review. Iran J Basic Med Sci. 2019; 22(3):225.

Nagpal I and Abraham SK. Ameliorative effects of gallic acid, quercetin and limonene on urethane-induced genotoxicity and oxidative stress in Drosophila melanogaster. Toxicol Mech Methods. 2017; 27(4):286- 292.

Mladenović M, Matić , tanić , olujić , Mihailović V, tanković N, atanić J. Combining molecular docking and 3-D pharmacophore generation to enclose the in vivo antigenotoxic activity of naturally occurring aromatic compounds: myricetin, quercetin, rutin, and rosmarinic

acid. Biochem Pharmacol. 2013; 86(9):1376-1396.




How to Cite

El-Taher, E. M. M., El-Sherei, M. M., El Dine, R. S., El Naggar, D. M. Y., Khalil, W. K. B., Eshak, M. G., … Kassem, M. E. S. (2020). Chemical Profiling of Acacia auriculiformis (A. Cunn.) ex Benth. Leaves Extract and its Impact on Gene Expression and Apoptotic DNA Damage in CCl4-Induced Hepatotoxicity in Male Rats: Tropical Journal of Natural Product Research (TJNPR), 4(11), 936–944. Retrieved from