Synthesis and Evaluation of Anticancer activity of some new
3(2h)-one pyridazinone derivatives by Molecular Docking Studies
Mehvish*, Arvind Kumar
SD College of Pharmacy and Vocational Studies, Muzaffarnagar, UP.
*Corresponding Author E-mail: mehvishakhlaq786pharma@gmail.com
ABSTRACT:
The present research aims to synthesize new series of 3(2h)-one pyridazinone derivatives with a potential anti-oxidant activity. All the compounds were synthesized by the esterification reaction of 2-(4- Fluoro Phenyl)--butyric acid with sulphuric acid and ethanol followed by hydrazine hydrate to give substituted hydrazides and then hydrazone with substituted ketons, which are cyclized with aldehyde substitutes to give the [5-(2,5-Dichloro-phenyl)-2-oxo-3,4-dihydro-2H-pyridin-1-yl]-acetic acid(4-4chloro-benzylidene)-hydrazide derivatives. Structure of all derivatives was confirmed by Fourier transform infrared spectroscopy (FT-IR) and Nuclear magnetic resonance (NMR) spectroscopy. All these compounds were demonstrated for in-vitro anti oxidant activity by using DPPH radical scavenging activity and hydrogen peroxide scavenging activity. Molecular Docking was carried out by cyclin dependent kinase protein and DNA-hexamer ATGCAT. Molecular docking studies of derivatives were carried out by AutoDock Vina molecular docking software. All the results were calculated by PaDEL software. Results of antioxidant activity revealed that all compounds are potent antioxidants at a concentration of 50µg/ml.
KEYWORDS: Pyridazinone, Molecular Docking, Anti-oxidant activity, NMR, FT-IR.
INTRODUCTION:
Antioxidant activity was determined by DPPH radical scavenging activity and hydrogen peroxide scavenging activity using ascorbic acid as standard5 and anticancer activity by molecular docking analysis on cyclin dependent kinase protein6 and DNA-hexamer ATGCAT 7. The molecular docking approach is widely used in the drug discovery process for the evaluation of the effective pharmacological activity of a particular drug by model the interaction of an active pharmaceutical component (API) and respective protein (receptor), at the atomic level, which allow us to examine the behavior of API in the binding site of receptor8,9,10. The docking process works by two basic steps: prediction of the ligand confirmation as well as its position and orientation within these sites (usually referred to as pose) and assessment of the binding affinity. Pyridazinone (Pyridazin-3(2H)-one) derivatives became a core of interest for pharmaceutical research on account of exhibiting several biological and pharmacological activities such as antihypertensive, diuretics, analgesic, anticancer and antithrombic. The nucleus of pyridazinone has been regarded as a wonder nucleus because of its pharmacological activities11,12,13. Pyradazinones are exploited as principle nucleus in several drugs such as Levosimendan, Pimobendan, Sulmazole, Indolidan, Milrinone, Emorfazone, Imazodan, Amipizone, and Zardaverine etc14,15,16. Pyradazinones derivatives contain two nitrogen atoms in a six member ring at 1 and 2 position and owned by an influential group of pyridazine heterocyclic compounds. In pyridazinone compounds both-carbonyl group and amine group are worthy placed with each other. Tautomerism has been reported in nearly all pyridazinone derivatives. Pyridazinones principally exist in oxo form as evaluated in its various derivatives17,18.
MATERIALS AND METHODS:
Methodology:
General method for the synthesis of pyridazin-3(2H)-one
Step-2 Synthesis of (5-(4-Fluoro-cyclohexyl)-hexa-1,5-dien-2-ol) from 2,2-(4-Fluoro Phenyl)-butyric acid:
(5-(4-Fluoro-cyclohexyl)-hexa-1,5-dien-2-ol was synthesized from their respective 2,2-(4-Fluoro Phenyl)-butyric acid, using excess of dry ethanol in the presence of sulphuric acid under reflux for 20-24 hours and identified by TLC in every 30 min.
Step-4 Synthesis of different derivatives of ([3-2,2fluoro-phenyl)-6-oxo-5,6-dihydro4H-pyridazin-1-yl]-aceticacid(4-fluoro-benzylidene)hydrazide i.e [(n1(a), (n1(b), (n1(c), (n1(d), (n1(e), (n2(a), (n2(b) and (n2(c)]: Synthesis of different derivatives of hydrazide was synthesized from R- 22-(4-Fluoro Phenyl)-butyric acid hydrazide with the addition of substituted ketone in ethanol and glacial acetic acid in 100ml round bottom flask by refluxing for 45 min and identified by TLC in every 30 min. After this the reaction mixture was poured in ice- cold water. The solid product was collected by filtration under suction. The product was recrystalized by using chloroform. All the compounds were analyzed by FT-IR and NMR spectroscopy.
Characterization:
1. Identification: All the synthesized compounds were identified by melting point analysis, FT-IR and NMR spectroscopy.
2. Determination of Anticancer activity by molecular docking
I. Data Base:
For molecular docking studies, National Center for Biotechnology Information’s (NCBI) website 19 and Protein Data Bank’s (PDB)20 website were used as biological and receptor sources. For designing and optimizing the geometry of the derivatives, Chemdraw Ultra 7.0 was used and molecular docking studies21 of derivatives were carried out by AutoDock Vina molecular docking software. All the results were calculated by PaDEL software22.
II. Molecular designing and optimization:
The chemical structures of the hybrid derivatives were drawn using Chem Draw Ultra 7.0 and energy minimization of derivatives was achieved with Chem3D Pro of chem office suit for taking energy of each molecule upto its lowest energy state (highest stability). 3D structure of 2H-Pyridazin-3one Complex protein (PDB id: 1X95 and 4MNB) was retrieved from PubChem compound database at NCBI.
III. Protein Preparation:
Wizard of Maestro software was used for protein preparation23. The protein structures, namely, 1X95 (n1) and 4MNB (n2) were taken from Protein Data Bank. The selected chains were edited for missing hydrogen and for assigning proper bond orders. The H-bonds were optimized using sample orientations.
The in vitro antioxidant activity was carried out at MBC Industries REF.NO:MBC/QC/2021-22/03 (A Pharma Manufacturer), Village: Johron, Trilokpur Road, Kala-Amb, Distt.-Sirmour (H.P.)-173030 following the below mentioned procedures by evaluating DPPH Radical Scavenging Activity and Hydrogen peroxide-scavenging activity.
I. DPPH Radical Scavenging Activity:
The activity is based on the scavenging of 1, 1-diphenyl-2-picrylhydrazyl free radical in stable form. DPPH activity of all synthesized compound was carried out by adding a different concentrations of all hydrazide derivatives in 3ml of 0.004% methanolic solution of 1, 1-diphenyl-2-picrylhydrazyl.Absorbance was determined at 517nm. Percentage inhibition was calculated using given equation after the addition of 30 minutes24-29.
% Inhibition = (A0-A1) / A0 x 100
Where, A0= absorbance of control and A1= absorbance of sample. Using this percentage inhibition reading the value of IC50 was calculated.
II. Hydrogen peroxide-scavenging activity:
The activity is based on the scavenging of hydrogen peroxide free radical. Hydrogen peroxide-scavenging activity was carried out by dissolving different concentrations of all hydrazide derivatives in 3.4ml of 0.1 M phosphate buffer. Absorbance was determined at 230nm at interval of 10 min against hydrogen peroxide as blank solution. % Inhibition was calculated using following equation30 - 34:
% Inhibition = [(A0-A1)/A0] × 100, Where A0 = the absorbance of the control, A1 = absorbance of the sample.
I. n1-(a) (N-[(E )-(chlorophenyl)methylidene]-2-[3-(4-fluorophenyl)-6oxo-5,6-dihydropyridazin-1(4H)-ylacetohydrazide:
Light brown colored flakes, m.p= 69-89 ºC, Rf= 0.56, %yield=90.00 %, IR (KBr, cm-1, ʋ): 3434.83(-NH-);3019.13(-CH-); 1655.02(-C=O); 1385.04(-N=C); 1215.52(-O-)1HNMR(CDCl3, 400 MHz): δ6.936-8.499 (m, 9H, Ar-H), 0.850-3.916 (m, 12H), 5.807 (s, 1H, NH);
II. n1-(b) (2-3[3-(4-fluorophenyl)-6oxo-5,6-dihydropyridazin-1(4H)-yl]-N-[(E)-4nitrophenyl)methylidene]acetohydrazide:
Light Brown colored flakes, m.p.=59-89ºC, Rf= 0.56, %yield=92.95 %, IR (KBr, cm-1, ʋ): 3334.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-),1HNMR(CDCl3, 400 MHz): δ 6.966-8.499 (m, 9H, Ar-H), 0.850-4.916 (m, 12H), 4.807 (s, 1H, NH);
III. n1-(c) (2-[3-(4-fluorophenyl)-6-oxo-5,6dihydropyridazin-1(4H)-yl]-N-(E )-(4-methoxyphenyl)methylidene]acetohydrazide:
Light yellow colored flakes, m. p.=50-99 ºC, Rf=0.56, % yield=98.95 %, IR (KBr, cm-1, ʋ): 3234.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-) 1HNMR(CDCl3, 400 MHz): δ 6.936-8.499 (m 9H, Ar-H), 0.880-3.916 (m,12H), 4.808 ( 1H, NH);
IV. n1-(d) N-[(4-Fluorophenyl)Methyl]-9-Hydroxy-2-(2-Morpholinoethyl)-1,8-Dioxo-Pyrido[1,2-A]Pyrazine-7-Carboxamide:
orange colored flakes, m. p=78-79ºC, Rf=0.66, % yield=78.95 %, IR (KBr, cm-1, ʋ): 2234.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-)1HNMR(CDCl3, 400 MHz): δ 8.456-8.499 (m, 9H, Ar-H), 0987-3.657 (m, 10H ), 4098 (s, 1H, NH);
V. n1-(e) (2-[3-(4-fluorophenyl)-6-5,6dihydropyridazin-1(4H)-yl]-N-[(E)-phenylmethylidene]acetohydrazide:
Orange colored flakes, m. p= 88-89 ºC, Rf=0.66, %yield=68.95 %, IR (KBr, cm-1, ʋ): 3234.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-) 1HNMR(CDCl3, 400 MHz): 56.876-8.880 (m, 9H, Ar-H), 0.850-3.916 (m, 9H), 8.909 (s, 1H, NH);
VI. n2-(a) N-[E)-4aminophenyl)methylidene]-2[3-(4-chloro-3-methylphenyl)-6oxo-5,6dihydropyridazine-1(4H)-yl]acetohydrazide:
Orange colored flakes, m. p.=57-67 ºC, Rf= 0.66, %yield=40.56%, IR (KBr, cm-1, ʋ): 3224.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-) 1HNMR(CDCl3, 400 MHz): δ 4.897-8.788 (m, 9H, Ar-H), 0.850-3.916 (m, 8H), 3.809 (s, 1H, NH);
VII. n2-(b) 6-(4-Chloro-3-methyl-phenyl)-2-{2-oxo-3-[(3,4,5-triamino-benzylidene)-amino]-propyl}-4,5-dihydro-2H-pyridazin-3-one:
Light yellow colored flakes, m. p=47-68 ºC, Rf=0.66, %yield=56.59 %, IR (KBr, cm-1, ʋ): 1224.83(-NH-); 3019.13(-CH-); 1535.02(-C=O); 1385.04(-N=C); 12415.52(-O-)1HNMR(CDCl3, 400 MHz): δ 6.939-8.567 (m 9H, Ar-H), 0.899-3.916 (m,12H), 4.789 ( 1H, NH);
VIII. n2-(c) [3-(4-Chloro-3-methyl-phenyl)-6-oxo-5,6dihydropyridazin-1(4H)-yl]-N-(4-nitrophenyl)methylidene}acetohydrazide:
Orange colored flakes, m. p.= 88-89 ºC, Rf =0.66, % yield=68.95 %, IR (KBr, cm-1, ʋ): 3234.83(-NH-); 3019.13(-CH-); 1635.02(-C=O); 1385.04(-N=C); 1215.52(-O-)1HNMR(CDCl3, 400 MHz): δ6.936-8.499 (m, 9H, Ar-H), 0.850-3..098 (m, 12H), 5.677 (s, 1H, NH);
2. Molecular docking studies:
All the results of the molecular docking study revealed a potent anticancer activity of all synthesized derivatives. All the polar hydrogen were displayed and compounds (n1and n2) showed marked binding selectivity with receptors of cyclin dependent kinase protein (PDB-4MNB)and intercalation with DNA-hexamer ATGCAT (PDB ID-1X95). SAR of these compounds demonstrated the existence of hetrocyclic ring electron withdrawing group on pyridazinone nucleus. Both of these revelations are correlated with excellent binding affinity and potency of compounds. Results are shown in and table no.1 and in figures (from 5 to 12) separately.
Figure 1: Ligand-interaction diagrams of representative compound no 1 (n1-(a)) with DNA (dt2(A) chain with this binding affinity -7.3 and this distance nitrogen(N4) to oxygen(O2) 3.07Å.
Figure 2: Ligand-interaction diagrams of representative compound no2 (n1-(b)) with DNA (dt2(A) chain with this binding affinity -7.4 and this distance nitrogen (N4) to oxygen (O2) 3.07Å
Figure 3: Ligand-interaction diagrams of representative compound no 3 (n1-(c)) with DNA (dt2(A) chain with this binding affinity -7.4 and this distance nitrogen (N4) to oxygen (O2) 3.17Å.
Figure 4: Ligand-interaction diagrams of representative compound no 4 (n1-(d)) with DNA(dt2(A) chain with this binding affinity -7.3 and this distance nitrogen(N4) to oxygen(O2) 3.24Å.
Figure 5: Ligand-interaction diagrams of representative compound no 5 (n1-(e) with DNA (dg3(A)) chain with this binding affinity -8.00 and this distance nitrogen (N4) to oxygen (O2) 3.00Å.
Figure 6: Ligand-interaction diagrams of representative compound no 11 (n2-(a) with DNA (dg3(A)) chain with this binding affinity -7.02 and this distance oxygen (O2) to nitrogen (N4) 3.12Å.
Figure 7: Ligand-interaction diagrams of representative compound no 12 (n2-(b) with DNA (dc4(A)) chain with this binding affinity -7.00 and this distance oxygen(O2) to nitrogen(N4) 2.93Å
Figure 8: Ligand-interaction diagrams of representative compound no 13 (n2-(c) with DNA(dt5(A)) chain with this binding affinity - 4.9 and this distance oxygen(O2) to nitrogen(N4) 2.47A
3. Anti oxidant activity:
The results of anti-oxidant activity by DPPH Radical Scavenging activity and hydrogen peroxide-scavenging activity of six compounds (n1-(a), n1-(b), n1-(c), n2-(a), n2-(b) and n2-(c) is shown in table 1. All results revealed the potent in-vitro anticancer activity in the concentration of 50µg/ml.
Table 1: Scavenging effects of compounds (n1-(a), n1-(b), n1-(c) and standard ascorbic acid on DPPH radical and H2O2
|
Concentration (µg/ml) |
Scavenging effects on DPPH radical |
Scavenging effects on H2O2
|
||||||
|
n1-(a) |
n1-(b) |
n1-(c) |
Ascorbic acid % inhibition |
n2-(a) |
n2-(b) |
n2-(c) |
Ascorbic acid % inhibition |
|
|
10 |
3.11 |
15.4 |
9.31 |
36.42 |
15.20 |
17.32 |
8.35 |
17.09 |
|
20 |
9.84 |
31.7 |
22.53 |
54.67 |
25.38 |
32.71 |
27.81 |
50.06 |
|
30 |
26.70 |
81.54 |
82.22 |
74.41 |
38.03 |
41.26 |
35.12 |
59.26 |
|
40 |
82.81 |
89.33 |
86.31 |
92.15 |
49.21 |
71.63 |
50.33 |
69.91 |
|
50 |
92.20 |
91.35 |
90. 62 |
94.13 |
53.67 |
82.67 |
56.85 |
76.24 |
|
IC50 (µg/ml) |
65 |
25 |
33 |
24 |
75 |
55 |
70 |
30 |
The results of in-vitro antioxidant studies are revealed using ANOVA accompanied by Dunnett test.
CONCLUSION:
The authors are thanks to animal house of S. D. College of Pharmacy and Vocational Studies, Muzaffarnagar to providing animals for pharmacological screening. Thanks are also due to IIT Roorkee for recording the IR and NMR spectra of the synthesized compounds and to MBC Industries, Kala-Amb, Himachal Pradesh for allowing authors to carry out antioxidant activity.
REFERENCE:
1. Pham-Huy LA, Hua H, Pham-Huy C. Free radicals, antioxidants in disease and health. Int J Biomedical Sci. 2008;4(2): 89–96.
2. Pisoschi AM, Pop A, Cimpeanu C, Predoi G. Antioxidant capacity determination in plants and plant derived products: A review. Oxidative Medicine and Cellular Longevity. 2016:36.
3. Gupta D. Methods for Determination of Antioxidant Capacity: A Review. Int J Pharm Sci Res. 2015; 6(2): 546-66. doi: 10.13040/IJPSR.0975-8232.6 (2).546-66.
4. Cem Y, Ozan G, Burcu k, Cobanoglu S, Sukuroglu K, Dogruer DS. Synthesis of some 3(2H) - pyridazinone and 1(2H) - phthalazinone derivatives incorporating aminothiazole moiety and investigation of their antioxidant, acetylcholinesterase, and butyrylcholinesterase inhibitory activities. Medicinal Chemistry Research. 2015;24:2010-2017. 10.1007/s00044-014-1205-8
5. Ali BM, Boothapandi M, Salam A, Nasar S. Nitric oxide, DPPH and hydrogen peroxide radical scavenging activity of TEMPO terminated polyurethane dendrimers: Data supporting antioxidant activity of radical dendrimers. Data in Brief. 2020;20:2352-3409. https://doi.org/10.1016/j.dib.2019.104972.
6. Sharma V, Sharma PC, Kumar V. In silico molecular docking analysis of natural pyridoacridines as anticancer agents. Advances in Chemistry. 2016:9.
7. Ricci GC, Netz PA. Docking Studies on DNA-Ligand Interactions: Building and Application of a Protocol To Identify the Binding Mode. Journal of Chemical Information and Modeling. 2009;49;1925-35. 10.1021/ci9001537.
8. Meng XY, Zhang HX, Mezei M, Cui M. Molecular docking: a powerful approach for structure-based drug discovery. Current Computer-aided Drug Design. 2011;7(2):146–157. https://doi.org/10.2174/157340911795677602
9. Acharya R, Chacko S, Bose, P, Lapenna A, Pattanayak SP. Structure Based Multitargeted Molecular Docking Analysis of Selected Furanocoumarins against Breast Cancer. Sci Rep. 2019;9:15743. https://doi.org/10.1038/s41598-019-52162-0
10. Mohd A, Vinay KM, Manish KD, Kashyap MM, Pratap S, Kumar KS, Sanmukh UR, Umamaheswari A, Singh S. In silico Prediction, Characterization, Molecular Docking, and Dynamic Studies on Fungal SDRs as Novel Targets for Searching Potential Fungicides Against Fusarium Wilt in Tomato. Frontiers in Pharmacology. 2018;9: 1038 DOI=10.3389/fphar.2018.01038
11. Seth S, Sharma A, Raj D. Pyridazinones: A wonder Nucleus with scaffold of Pharmacological activities. American journal of biological and Pharmaceutical Research. American Journal of Biological and Pharmaceutical Research. 2014;(1(3):105-116.
12. Asif M, Deewan S, Anita S. Analgesic Activity of Some 6-Phenyl-4-Substituted Benzylidene Tetrahydro Pyridazin-3(2H)-Ones. Global Journal of Pharmacology. 2011;(1(3):18-22.
13. Asif M, Anita S. Anticonvulsant Activity of 4-(Substituted Benzylidene)-6-(3-nitrophenyl)-4,5-dihydro Pyridazin-3(2h)-ones Against Maximal Electro Shock Induced Seizure. Middle-East Journal of Scientific Research. 2011;9(4):481-485.
14. Seth S, Sharma A, Raj D. Synthesis of new Mannich bases of arylpyridazinones as analgesic and anti- inflammatory agents. American Journal of Biological and Pharmaceutical Research. 2014;1(3):105-116.
15. Guan LP, Sui X, Deng XQ, Quan YC, Quan ZS. Synthesis and anticonvulsant activity of a new 6-alkoxy-[1,2,4]triazolo[4,3- b]pyridazine. Eur J. Med. Chem. vol.2010;45(3):1746-1752.
16. Fang-Zong Hu, Zhang GF, Zhu YQ, Zou XM, Liu B, Yang HZ. Synthesis and biological activity of 3-aryloxy-6-(3,5-dimethyl-1H-pyrazol-1-yl)-pyridazine. Chin. J. Org. Chem.,vol. 2007;27:758-762.
17. Kawano YH, Nagayaand M, Gyoten. Preparation of fused pyridazine compounds as drugs. WO Patent 2000020417. Chem.Abstr. 2000;132:265202.
18. Rong DL, Zhai X, Zhao YF, Yu S, Gong P. Synthesis and anti-tumor activities of a novel series of tricyclic 1-anilino-5H-pyridazino[4,5-b]indoles. Arch. Pharm. 2007;340:424-428.
19. NCBI Resource Coordinators. Database resources of the National Center for Biotechnology Information. Nucleic acids research. 41(Database issue). 2-013 D8–D20. https://doi.org/10.1093/nar/gks1189
20. Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, Weissig H, Shindyalov IN, Bourne PE. The Protein Data Bank. Nucleic acids research. 2000;28(1):235–242. https://doi.org/10.1093/nar/28.1.235
21. Ajeet . In silico designing and characterization of Amiloride derivatives as ion channel modulator. Med Chem Res. 2013;22: 1004–1010. https://doi.org/10.1007/s00044-012-0096-9
22. Mahmud AW, Shallangwa GA, Uzairu A. QSAR and molecular docking studies of 1,3-dioxoisoindoline-4-aminoquinolines as potent antiplasmodium hybrid compounds. Heliyon. 2020;6(3). https://doi.org/10.1016/j.heliyon.2020.e03449.
23. Garikapati MS, Matvey DA, Day T, Annabhimoju R, Woody S. Protein and ligand preparation: Parameters, protocols, and influence on virtual screening enrichments. Journal of Computer-Aided Molecular Design. 2013; 27(3). 10.1007/s10822-013-9644-8.
24. Sharma US, Kumar A. In vitro antioxidant activity of Rubus ellipticus fruits. Journal of Advanced Pharmaceutical Technology and Research. 2011;2(1):47–50. https://doi.org/10.4103/2231-4040.79805
25. Rahman MM, Islam MB, Biswas M, Alam AHM. In vitro antioxidant and free radical scavenging activity of different parts of Tabebuia pallida growing in Bangladesh. BMC Res Notes. 2015; 8(621). https://doi.org/10.1186/s13104-015-1618-6.
26. Amini MH, Kalsi V, Kaur B, Gopal LK, Lobo R, Singh G, Agarhari UC, Yele S, Suttee A. Phytochemical screening and antioxidant activity of Heracleum afghanicum kitamura leaves. Research J. Pharm. Tech. 2017;10(10):3498-3502.
27. Som KM, Manik S, Javaid I, Mohd Y, Razda S. Phytochemical analysis, total flavonoids, phenolic contents and antioxidant activity of extract from the leaves of Rhododendron arboretum. Research J. Pharm. Tech. 2020;13(14):1193-1197.
28. Choudhary G, Choudhary V, Baheti A, Mantri M, Matapurkar S, More C. Synergistic antioxidant activity of a polyherbal preparation. Research J. Pharm. And Tech. 2020;13(3):1193-1197.
29. Pal R, Girhepunje K, Shrivastva N, Hussain MM, Thirumoorthy N. Antioxidant and free radical scavenging acrivity of ethanolic extract of Morinda citrifolia. Research J. Pharm. And Tech. 2011;4(8):1224-1226.
30. Suryawanshi A, Chougule P, Jain A. In-vitro antioxidant activity of hexanolic extract of trachypermum ammi. Research J. Pharm. And Tech. 2015;8(10):1403-1408.
31. Joshi H, Pagare M, Patil L, Kadam V. In-vitro antioxidant activity of ethanolic extract of leaves of Buchanania Lanzan Spreng. Research J. Pharm. And Tech. 2011;4(6):920-924.
32. Sharma S, Pal A. Evaluation of in vitro antioxidant properties of methanol and aqueous extracts of Parkinsonia aculeate L. Leaves. The Scientific World Journal. 2013;7.
33. GÜLÇin I, Alici HA, Cesur M. Determination of in vitro antioxidant and radical scavenging activities of propofol. Chemical and Pharmaceutical Bulletin. 2005;53(3):281-5.
34. Nabavi SM, Ebrahimzadeh MA, Nabavi SF, Fazelian M, Eslami B. In vitro antioxidant and free radical scavenging activity of Diospyros lotus and Pyrus boissieriana growing in Iran. Pharmacognosy Magazine. 2009;5(18):122.
Received on 02.03.2021 Modified on 25.04.2021
Accepted on 28.05.2021 © RJPT All right reserved
Research J. Pharm. and Tech. 2022; 15(5):2279-2284.
DOI: 10.52711/0974-360X.2022.00379