Abstract
Inflammation and angiogenesis are two major contributors to tumourigenesis. Melilotus indicus is traditionally used as an anti-inflammatory agent. The current study was designed to investigate the anti-inflammatory and anti-angiogenic properties of ethanolic extract of M. indicus (Miet) whole plant and its marker compound (coumarin) using a series of in vivo methods. Extraction by maceration was adopted to prepare ethanolic extract. Phytochemical compounds present in Miet were investigated using both qualitative and quantitative methods. In vivo safety profile of Miet was investigated in behavioural studies. Four acute oedema models such as carrageenan, serotonin, histamine-induced paw oedema and xylene-induced ear oedema, and chronic formaldehyde-induced paw oedema model were employed to explore the anti-inflammatory potential of Miet. Chorioallantoic chick membrane assay (CAM) was performed to explore anti-angiogenic potential of Miet. Histopathological evaluations were conducted to access improvement in skin texture of paws. TNF-α ELISA kit was used to study effects of treatment on serum levels of TNF-α. Extraction by maceration resulted in formation of greenish coloured semisolid extract with a high coumarin content. In vivo toxicological studies revealed LD50 of Miet was greater than 8000 mg/kg. Data of acute inflammatory models depicted significant (p < 0.05) inhibition of oedema in Miet, coumarin and standard (piroxicam/indomethacin) treated groups. 750 mg/kg of Miet induced comparable (p > 0.05) anti-inflammatory effects to that of standard-treated groups. Coumarin showed better anti-inflammatory effects in carrageenan-induced paw oedema model as compared with histamine- and serotonin-induced oedema models. Data of chronic inflammatory models also depicted dose-dependent anti-inflammatory attributes of Miet which were comparable with standard treated groups. Significant (p > 0.05) downregulation of TNF-α in serum samples of animals treated with Miet and piroxicam was observed as compared with control group. Furthermore, Miet significantly halted blood vessels formation in CAM assay. Overall, data of the current study highlight that M. indicus has anti-inflammatory and anti-angiogenic potentials, and, thus, can potentially be used as an adjuvant therapy in solid tumours management.
Similar content being viewed by others
References
Ahmed D, Baig H, Zara S (2013) Seasonal variation of phenolics, flavonoids, antioxidant and lipid peroxidation inhibitory activity of methanolic extract of Melilotus indicus and its sub-fractions in different solvents. Int J Phytomedicine 4:326–332
Akindele A, Adeyemi O (2007) Antiinflammatory activity of the aqueous leaf extract of Byrsocarpus coccineus. Fitoterapia 78:25–28
Amdekar S, Roy P, Singh V, Kumar A, Singh R, Sharma P (2012) Anti-inflammatory activity of lactobacillus on carrageenan-induced paw edema in male wistar rats. Int J Inflam 2012:752015
Asif M et al (2016a) Isoledene from Mesua ferrea oleo-gum resin induces apoptosis in HCT 116 cells through ROS-mediated modulation of multiple proteins in the apoptotic pathways: a mechanistic study. Toxicol Lett 257:84–96
Asif M et al (2016b) Anticancer attributes of Illicium verum essential oils against colon cancer. South Afr J Botany 103:156–161
Asif M et al (2020) Evaluation of in vivo anti-inflammatory and anti-angiogenic attributes of methanolic extract of Launaea spinosa. Inflammopharmacol. https://doi.org/10.1007/s10787-020-00687-6
Baci D et al (2019) Downregulation of pro-inflammatory and pro-angiogenic pathways in prostate cancer cells by a polyphenol-rich extract from olive mill wastewater. Int J Mol Sci 20:307
Balkwill F, Mantovani A (2001) Inflammation and cancer: back to Virchow? Lancet 357:539–545
Braga PC et al (2013) Antioxidant activity of Melilotus officinalis extract investigated by means of the radical scavenging activity, the chemiluminescence of human neutrophil bursts and lipoperoxidation assay. J Med Plant Res 7:358–365
Bukhari IH, Ramzan M, Riaz M, Bokhari TH, Munir S (2013) Determination of trace heavy metals in different varieties of vegetables and fruits available in local market of Shorkot Pakistan. Int J Curr Pharm Res 5:101–105
Cursiefen C et al (2004) VEGF-A stimulates lymphangiogenesis and hemangiogenesis in inflammatory neovascularization via macrophage recruitment. J Clin Investig 113:1040–1050. https://doi.org/10.1172/JCI20465
Day RO, Graham GG (2013) Non-steroidal anti-inflammatory drugs (NSAIDs). BMJ 346:f3195
Deryugina EI, Quigley JP (2015) Tumor angiogenesis: MMP-mediated induction of intravasation-and metastasis-sustaining neovasculature. Matrix Biol 44:94–112
Dghaim R, Al Khatib S, Rasool H, Ali Khan M (2015) Determination of heavy metals concentration in traditional herbs commonly consumed in the United Arab Emirates. J Environ Public Health 2015:973878
Ginwala R, Bhavsar R, Chigbu DI, Jain P, Khan ZK (2019) Potential role of flavonoids in treating chronic inflammatory diseases with a special focus on the anti-inflammatory activity of apigenin. Antioxidants 8:35
Gul R, Jan SU, Faridullah S, Sherani S, Jahan N (2017) Preliminary phytochemical screening, quantitative analysis of alkaloids, and antioxidant activity of crude plant extracts from Ephedra intermedia indigenous to Balochistan. Sci World J 2017:5873648
Hadjipavlou-Litina D, Litinas K, Christos K (2007) The anti-inflammatory effect of coumarin and its derivatives. Antiinflamm Antiallergy Agents Med Chem 6:293–306
Hanahan D, Weinberg R (2011) Hallmarks of cancer: the next generation. Cell 144:646–674
Hosseinzadeh H, Ramezani M, Salmani G-A (2000) Antinociceptive, anti-inflammatory and acute toxicity effects of Zataria multiflora Boiss extracts in mice and rats. J Ethnopharmacol 73:379–385
Hussain T, Tan B, Yin Y, Blachier F, Tossou MCB, Rahu N (2016) Oxidative stress and inflammation: what polyphenols can do for us? Oxidat Med Cell Long 2016:7432797
Joy A, Appavoo M, Mohesh M (2016) Antiangiogenic activity of Strychnos nux vomica leaf extract on chick chorioallantoic membrane model. J Chem Parm Res 8:549–552
Kubrak T, Podgórski R, Stompor M (2017) Natural and synthetic coumarins and their pharmacological activity. Eur J Clin Exp Med 15:169–175
Kumar T, Jain V (2014) Antinociceptive and anti-inflammatory activities of Bridelia retusa methanolic fruit extract in experimental animals. Sci World J 2014:890151
Kunle O, Folashade O, Egharevba HO, Omoregie H, Ahmadu PO (2012) Standardization of herbal medicines: a review. Int J Biodiv Conserv 4:101–112
Liu FT, Rabinovich GA (2010) Galectins: regulators of acute and chronic inflammation. Ann N Y Acad Sci 1183:158–182
Maiga A, Diallo D, Bye R, Paulsen BS (2005) Determination of some toxic and essential metal ions in medicinal and edible plants from Mali. J Agric Food Chem 53:2316–2321. https://doi.org/10.1021/jf040436o
Mantovani A, Garlanda C, Allavena P (2010) Molecular pathways and targets in cancer-related inflammation. Ann Med 42:161–170
Misra D, Mandal M, Ghosh NN, Mandal V (2018) Pharmacognostic standardization of an ethnomedicinal aquatic herb, Monochoria hastata (L.) Solms for its antibacterial potentiality. Pharmacogn J 10:533–540
Njoku VO, Obi C (2009) Phytochemical constituents of some selected medicinal plants. Afr J Pure Appl Chem 3:228–233
Paracatu LC, Zeraik ML, Bertozo LC, Bartolomeu AA, Filho L, Fonseca LMD, Ximenes VF (2016) Synthesis, antioxidant and anti-inflammatory properties of an apocynin- derived dihydrocoumarin. Med Chem 13:93–100
Pérez-Cano F, Castell M (2016) Flavonoids, inflammation and immune system. Nutrients 8:659
Reuter S, Gupta SC, Chaturvedi MM, Aggarwal BB (2010) Oxidative stress, inflammation, and cancer: how are they linked? Free Radical Biol Med 49:1603–1616
Saleem U, Hussain K, Ahmad M, Irfan Bukhari N, Malik A, Ahmad B (2014) Physicochemical and phytochemical analysis of Euphorbia helioscopia (L.). Pakistan J Pharm Sci 2014:27
Sethi G, Sung B, Aggarwal BB (2008) TNF: a master switch for inflammation to cancer. Front Biosci 13:5094–5107
Storgard C, Mikolon D, Stupack DG (2005) Angiogenesis assays in the chick CAM. Methods Mol Biol 294:123–136
Witaicenis A et al (2014) Antioxidant and intestinal anti-inflammatory effects of plant-derived coumarin derivatives. Phytomedicine 21:240–246
Yadav R, Agarwala M (2011) Phytochemical analysis of some medicinal plants. J Phytol 3:10–14
Yong YK, Sulaiman N, Hakim MN, Lian GEC, Zakaria ZA, Othman F, Ahmad Z (2013) Suppressions of serotonin-induced increased vascular permeability and leukocyte infiltration by Bixa orellana leaf extract. Biomed Res Int 2013:463145
Zhao J, Maitituersun A, Li C, Li Q, Xu F, Liu T (2018) Evaluation on analgesic and anti-inflammatory activities of total flavonoids from Juniperus sabina. Evidence-Based Complement Altern Med 2018:7965306
Acknowledgements
We would like to thank Faculty of Pharmaceutical Sciences, Government College University Faisalabad. This work was financially supported by the Higher Education Commission of Pakistan (Research Grant No: 21-1828/SRGP/R&D/HEC/2018). We are thankful to Dr Sven Peterson for proofreading the article.
Author information
Authors and Affiliations
Corresponding authors
Ethics declarations
Conflict of interest
Authors do not have any a conflict of interest in the current work. Mohammad Saleem, Muhammad Asif, Anum Parveen, Jawaria Asif, Hafiza Sidra Yaseen, Malik Saadullah and Mehwish Arif contributed equally in the current work.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Saleem, M., Asif, M., Parveen, A. et al. Investigation of in vivo anti-inflammatory and anti-angiogenic attributes of coumarin-rich ethanolic extract of Melilotus indicus. Inflammopharmacol 29, 281–293 (2021). https://doi.org/10.1007/s10787-020-00703-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10787-020-00703-9