Screening of secondary metabolites and immunostimulation potential of poly herbal formoulation (Asparagus racemosus, Mucuna pruriens, Anacyclus pyrethrum and Tribulus terristeris) in Wistar Albino rats

M. Akram, Shifa shaffique



Immune system is a defensive mechanism system which is responsible to defend against several foreign invading agents that harm individual health. It generates several types of cells which have ability to recognize, eliminate and kill all the undesirable agents. The immune system works by inhibition, multiplication, amplification, expression and induction response at different phases of immune system. Hence immunology is the most advance and recent biomedical research system which covers the prevention and treatment of wide range of medical disorders.  In this study, Immunomodulatory activity of Polyherbal formulation was determined. First of all, plants methanolic extracts of Asparagus racemosus, Mucuna pruriens, Anacyclus pyrethrum, Tribulus terrestris were prepared. The methanolic extracts of each plant were taken separately in equal amount and mixed to prepare the Polyherbal formulation (PHF). Screening test was applied on PHF for quantitative analysis. Hemolytic activity, thrombolytic activity and antioxidant activity were determined on experimental animals. To demonstrate the effect of standard Immunol drug, CCl4 and polyherbal formulation extract on hematological parameters. The methanolic extract of PHF was dissolved in distilled water and the dose was selected at 100, 200 and 300 mg/kg body weight. Blood sample collected in EDTA as anticoagulant were processed to determine the effect of PHP on selected hematological parameters including hemoglobin (Hb), red blood cell counts (RBCs), white blood cells (WBCs) count and Platelets count. Liver enzymes like transaminases (ALT and AST), uric acid, urea and creatinine were determined by automated photometric method to evaluate the drug safety. Serum samples were using for the estimation of Immunoglobulin M and Immunoglobulin G using enzyme linked immunosorbent assay (ELISA). Further, the C-reactive protein (CRP) was also estimated using standard protocol. Phytochemical screening of methanolic extract of polyherbal formulation showed the presence of alkaloids, tannins, flavonoids, phenols and cardiac glycosides. Bioactive constituent’s alkaloids, flavonoids and phenols were present in higher concentration. The maximum hemolysis was seen at a dose of 800μg/ml (40±0.38). The methanolic extracts of PHF were investigated for its thrombolytic activity. Addition of 800μl of streptokinase as positive control (30,000 IU/ ml) to clots and successive incubation for 90 minutes at 37 ºC showed 74.53±1.11% lysis of clot.  Distilled water was considered as negative control which exhibit insignificant percentage of lysis of clot (3.74±0.51). In the study, the methanolic extract of PHF showed highest thrombolytic activity at 800μg/ml concentrations (41±0.38) when compared with streptokinase (positive control). When all the groups were compared the serum level ALT, AST, TAS, TOS and OSI 50 index, a statistically significant difference was observed between the groups. A variety of antioxidant function approaches have been suggested. During the study, it was found that induction of CCl4 increases the number of altered macrophages. However, the administration of Polyherbal extract in CCl4 intoxicated animals, the number of altered macrophages came to level. Administration of polyherbal formulation extract in control mice also showed significant reduction of changed macrophages. Immunomodulatory drugs are substances that can alter the immune function of an individual, are called immunostimulants if their immune responses are enhanced, or are called immunosuppressant’s if they decrease. These medications are most widely used in infections, allergies and cancer infections. Modern medical treatment is also far-reaching because of the economic limitations in developed countries including Pakistan. For immunomodular operations, just a few plants have been tested. The analysis indicates that many medicinal plants and marine products have immunomodulatory function, however there is little proof that their use in clinical practice is not allowed. In future work in herbal medicine, therefore, immunomodulatory agents should become more relevant.


poly herbal formoulation,immune,stimulant.


of Pharmacy Research, 3 (2010), 1160-62. 4 Rosa M Aparicio, M José García-Celma, M Pilar Vinardell, and Montserrat Mitjans, 'In Vitro Studies of the Hemolytic Activity of Microemulsions in Human Erythrocytes', Journal of pharmaceutical and biomedical analysis, 39 (2005), 1063-67. 5 Mohammed Asad, Koteshwar Prasad, Licto Thomas, and Jagadish V Kamath, 'Evaluation of Anti-Arthritic and Anti-Inflammatory Activity of Sudard, a Poly Herbal Formulation', Iranian Journal of Pharmacology and Therapeutics, 6 (2007), 71-0. 6 K Sahira Banu, and L Cathrine, 'General Techniques Involved in Phytochemical Analysis', International Journal of Advanced Research in Chemical Science, 2 (2015), 25-32. 7 MEVLÜT Baskol, K Dolbun Seckin, and Gulden Baskol, 'Advanced Oxidation Protein Products, Total Thiol Levels and Total Oxidant/Antioxidant Status in Patients with Nash', Turk J Gastroenterol, 25 (2014), 32-7. 8 A Bhattacharya, AV Muruganandam, Vikas Kumar, and SK Bhattacharya, 'Effect of Poly Herbal Formulation, Eumil, on Neurochemical Perturbations Induced by Chronic Stress', (2002). 9 Guy A Cabral, and A Staab, 'Effects on the Immune System', Cannabinoids (2005), 385-423. 10 Ranjit Kumar Chandra, 'Nutrition and the Immune System: An Introduction', The American journal of clinical nutrition, 66 (1997), 460S-63S. 11 Saheli Chatterjee, S Chatterjee, KK Dey, and S Dutta, 'Study of Antioxidant Activity and Immune Stimulating Potency of the Ethnomedicinal Plant, Cassia Alata (L.) Roxb', Med. Aromat. Plants, 2 (2013). 12 Jiong-Ran Chen, Zhi-Qiang Yang, Ting-Jun Hu, Zuo-Ting Yan, Ting-Xian Niu, Ling Wang, Dong-An Cui, and Meng Wang, 'Immunomodulatory Activity in Vitro and in Vivo of Polysaccharide from Potentilla Anserina', Fitoterapia, 81 (2010), 1117-24. 13 Marco Colonna, 'Trems in the Immune System and Beyond', Nature Reviews Immunology, 3 (2003), 445-53. 14 Brian Cunningham, Peter Li, Bo Lin, and Jane Pepper, 'Colorimetric Resonant Reflection as a Direct Biochemical Assay Technique', Sensors and Actuators B: Chemical, 81 (2002), 316-28. 15 Mammen Daniel, Medicinal Plants: Chemistry and PropertiesScience publishers, 2006). 16 Peter J Delves, and Ivan M Roitt, 'The Immune System', New England journal of medicine, 343 (2000), 37-49. 17 Ozcan Erel, 'A Novel Automated Direct Measurement Method for Total Antioxidant Capacity Using a New Generation, More Stable Abts Radical Cation', Clinical biochemistry, 37 (2004), 277-85. 18 Kalpesh Gaur, ML Kori, and RK Nema, 'Comparative Screening of Immunomodulatory Activity of Hydro-Alcoholic Extract of Hibiscus Rosa Sinensis Linn. And Ethanolic Extract of Cleome Gynandra Linn', Global J Pharmacol, 3 (2009), 85-89. 19 Edoardo G Giannini, Roberto Testa, and Vincenzo Savarino, 'Liver Enzyme Alteration: A Guide for Clinicians', Cmaj, 172 (2005), 367-79. 20 Malgorzata Gorska-Ciebiada, Malgorzata Saryusz-Wolska, Anna Borkowska, Maciej Ciebiada, and Jerzy Loba, 'Serum Levels of Inflammatory Markers in Depressed Elderly Patients with Diabetes and Mild Cognitive Impairment', PloS one, 10 (2015), e0120433. 21 Shivaraj Gowda, Prakash B Desai, Vinayak V Hull, Avinash A K Math, Sonal N Vernekar, and Shruthi S Kulkarni, 'A Review on Laboratory Liver Function Tests', The Pan african medical journal, 3 (2009). 22 Temple Grandin, 'Animal Handling', Veterinary Clinics of North America: Food Animal Practice, 3 (1987), 323-38. 23 ———, 'Recommended Animal Handling Guidelines and Audit Guide', A systematic approach to animal welfare. American Industry Foundation: USA (2010). 24 Ankit Gupta, Madhu Naraniwal, and Vijay Kothari, 'Modern Extraction Methods for Preparation of Bioactive Plant Extracts', International journal of applied and natural sciences, 1 (2012), 8-26. 25 Asheesh Gupta, Nitin K Upadhyay, RC Sawhney, and Ratan Kumar, 'A Poly‐Herbal Formulation Accelerates Normal and Impaired Diabetic Wound Healing', Wound repair and regeneration, 16 (2008), 784-90. 26 Sandra Henkelman, Gerhard Rakhorst, John Blanton, and Willem van Oeveren, 'Standardization of Incubation Conditions for Hemolysis Testing of Biomaterials', Materials Science and Engineering: C, 29 (2009), 1650-54. 27 Maurice M Iwu, 'Handbook of African Medicinal Plants', (2014). 28 Eugène HJM Jansen, and Tatjana Ruskovska, 'Comparative Analysis of Serum (Anti) Oxidative Status Parаmeters in Healthy Persons', International Journal of Molecular Sciences, 14 (2013), 6106-15. 29 Niels Kaj Jerne, 'The Immune System', Scientific American, 229 (1973), 52-63. 30 Špela Magister, and Janko Kos, 'Cystatins in Immune System', Journal of Cancer, 4 (2013), 45. 31 Takashi Makinodan, and Marguerite MB Kay, 'Age Influence on the Immune System', Advances in immunology, 29 (1980), 287-330. 32 Ashis K Mukherjee, Sudhir K Rai, Rupamoni Thakur, Pronobesh Chattopadhyay, and Santosh K Kar, 'Bafibrinase: A Non-Toxic, Non-Hemorrhagic, Direct-Acting Fibrinolytic Serine Protease from Bacillus Sp. Strain as-S20-I Exhibits in Vivo Anticoagulant Activity and Thrombolytic Potency', Biochimie, 94 (2012), 1300-08. 33 Ralph Panstruga, Jane E Parker, and Paul Schulze-Lefert, 'Snapshot: Plant Immune Response Pathways', Cell, 136 (2009), 978. e1-78. e3. 34 Peter Parham, The Immune SystemGarland Science, 2014). 35 Jara Pérez-Jiménez, Sara Arranz, Maria Tabernero, M Elena Díaz-Rubio, José Serrano, Isabel Goni, and Fulgencio Saura-Calixto, 'Updated Methodology to Determine Antioxidant Capacity in Plant Foods, Oils and Beverages: Extraction, Measurement and Expression of Results', Food Research International, 41 (2008), 274-85. 36 Sweta Prasad, Rajpal Singh Kashyap, Jayant Y Deopujari, Hemant J Purohit, Girdhar M Taori, and Hatim F Daginawala, 'Effect of Fagonia Arabica (Dhamasa) on in Vitro Thrombolysis', BMC Complementary and Alternative Medicine, 7 (2007), 1-6. 37 Patrícia Rijo, Pedro L Falé, M Luísa Serralheiro, M Fátima Simões, Ana Gomes, and Catarina Reis, 'Optimization of Medicinal Plant Extraction Methods and Their Encapsulation through Extrusion Technology', Measurement, 58 (2014), 249-55. 38 Paola Sacerdote, 'Opioids and the Immune System', Palliative medicine, 20 (2006), 9-15. 39 William D Sawyer, Anthony P Fletcher, Norma Alkjaersig, and Sol Sherry, 'Studies on the Thrombolytic Activity of Human Plasma', The Journal of clinical investigation, 39 (1960), 426-34. 40 Pallavi Sharma, Ambuj Bhushan Jha, Rama Shanker Dubey, and Mohammad Pessarakli, 'Reactive Oxygen Species, Oxidative Damage, and Antioxidative Defense Mechanism in Plants under Stressful Conditions', Journal of botany, 2012 (2012). 41 Lauren M Sompayrac, How the Immune System WorksJohn Wiley & Sons, 2019). 42 Ben-Erik Van Wyk, and Michael Wink, Medicinal Plants of the WorldCABI, 2018). 43 Dhiraj A Vattem, Reza Ghaedian, and Kalidas Shetty, 'Enhancing Health Benefits of Berries through Phenolic Antioxidant Enrichment: Focus on Cranberry', Asia Pacific journal of clinical nutrition, 14 (2005), 120. 44 José A Vega, Olivia García‐Suárez, Jonas Hannestad, Marta Pérez‐Pérez, and Antonino Germanà, 'Neurotrophins and the Immune System', Journal of anatomy, 203 (2003), 1-19. 45 D Vrushali, and I Madhavi, 'Immunostimulatory Activity of Amoora Rohituka and Azadirachta Indica', Adv Pharmacol Toxicol, 7 (2006), 5-12. 46 Monika Waksmundzka-Hajnos, and Joseph Sherma, High Performance Liquid Chromatography in Phytoch


  • There are currently no refbacks.

Copyright (c) 2021 shifa shaffique

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

                AR Journals

Street 1st, Gaytri Vihar, Pinto Park, Gwalior, M.P. India

     (Design) 2009-2021


Follow @arjournals on Twitter