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Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity
Materials Today: Proceedings xxx (xxxx) xxx
Contents lists available at ScienceDirect
Materials Today: Proceedings journal homepage: www.elsevier.com/locate/matpr
Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity K. Pavithra 1, V.V. Sathibabu Uddandrao 1, S. Mathavan, N. Gobeeswaran, S. Vadivukkarasi, Saravanan Ganapathy ⇑ Centre for Biological Sciences, Department of Biochemistry, K. S. Rangasamy College of Arts and Science (Autonomous), Tiruchengode, Namakkal District, Tamilnadu 637215, India
a r t i c l e
i n f o
Article history: Received 7 April 2019 Received in revised form 9 May 2019 Accepted 13 May 2019 Available online xxxx Keywords: Abrus precatorius Bioactive compounds Antioxidants Medicinal plants Natural products
a b s t r a c t The current study was designed to develop fingerprint of the different bioactive compounds of Abrus precatorius and evaluation of its antioxidant activity. C-13 and H-NMR analysis of A. precatorius elucidated the key bioactive compounds in the leaves that were consistent with the classes of bioactive compounds detected by GC–MS analysis which revealed the presence of 9 major compounds. A. precatorius shown significant free radical scavenging activities and contain antioxidative enzymes which can regulate the free radical activity. In conclusion, the present study identified an array of bioactive compounds present in A. precatorius and reported their ethno-botanical uses against free radicals. Ó 2019 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Emerging Materials and Modeling.
1. Introduction Plants are an imperative source of bioactive molecules for drug discovery. Isolated bioactive molecules serve as starting materials for laboratory synthesis of drugs as well as a model for the production of biologically active compounds [1]. The search for natural bioactive compounds with prospective for the treatment and prevention of human diseases and to meet other needs is presently a key topic in many laboratories and industries. These compounds competently interact with DNA, proteins and other biological molecules to produce a desired outcome, which could be exploited for designing natural products-derived beneficial agents [2]. The study of medicinal plants starts with the pre-extraction and the extraction procedures, which is an important step in the processing of the bioactive constituents from plant materials. Considering their significant health effects, the proficient extraction methods of natural antioxidants, suitable assessment of antioxidant activity as well as their major resources from medicinal and food plants are depict enormous attention in food science and nutrition [3]. Plant-derived antioxidants are a large group of natural products with reducing or radical-scavenging capacity [4]. Due to their potent defensive, as well as medicinal actions, these compounds ⇑ Corresponding author. 1
E-mail address: [email protected] (S. Ganapathy). Both authors contributed equally to this work.
receive a great deal of attention by not only scientists but also pharmacologists and physicians [5]. In present time, medicinal plants as rich source of natural bioactive components are given precedence to study their antioxidant activity and discover their utilization in treatment of diabetes mellitus, dyslipidemia and cardiovascular diseases [6]. Abrus precatorius (Fabaceae) which is an ornamental, twining and woody vine originally native to India is now commonly found throughout the subtropical and tropical parts of the world [7]. In view of the above literature, we made an attempt to extract and identify the bioactive factors from A. precatorius by GC–MS and NMR and evaluated its antioxidant capability through in vitro studies.
2. Materials and methods 2.1. Sample preparation The fresh leaves of A. precatorius were collected from Tamilnadu, India and the plant was authenticated by Botanical Survey of India (BSI/SRC/5/23/2017/Tech/2874). The leaves were shade dried at 7 days and pulverized to powder in a mechanical grinder and extracted by continuous extraction using soxhlet apparatus with methanol for 72 h. The extract was collected and solvents were evaporated and dried in hot air oven at 50 °C–100 °C. The dried powder was collected and stored for further analysis.
https://doi.org/10.1016/j.matpr.2019.05.417 2214-7853/Ó 2019 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Emerging Materials and Modeling.
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
2
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
2.2. Preliminary phytochemical analysis The A. precatorius extract was subjected to qualitative examination for different phytoconstituents as per standard procedures [8]. 2.3. Nuclear magnetic resonance (NMR) analysis Hydrogen-NMR spectra of the extracts was performed using Bruker Biospin Avance 400-MHz NMR spectrophotometer with a 5 mm broad inverse probe head, equipped with shielded z-gradient accessories. Hydrogen-NMR spectral analysis of the extract was carried out using one-pulse sequence by dissolving the extract in 500 lL of deuterated methanol in 5 mm NMR tubes. A re-usable, sealed capillary tube containing 30 lL of 0.375% of trisodium phosphate in deuterium oxide was inserted into the NMR tube before recording the spectra. Tri-sodium phosphate served as chemical shift reference as well as internal standard for quantitative estimation. C-13 NMR heteronuclear single quantum correction was carried out using the Brucker’s standard pulse library [9]. 2.4. Gas chromatography-mass spectrometry (GC–MS) analysis The Clarus 680 GC was used in the analysis employed a fused silica column, packed with Elite-5MS (5% biphenyl 95% dimethylpolysiloxane, 30 m X 0.25 mm ID X 250 lm df) and the components were separated using Helium as carrier gas at a constant flow of 1 mL/min. The spectrums of the components were compared with the database of spectrum of known components stored in the GC–MS National Institute Standard and Technology (2008) library [10]. 2.5. Determination of in vitro antioxidant activity We assessed the antioxidant activity of A. precatorius extract by 2, 20 -diphenyl-1- picrylhydrazyl (DPPH) radical scavenging assay [11], 2,20 -azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging activity [12], activities of catalase (CAT) [13], glutathione peroxidase (GPx) [14] and superoxide dismutase (SOD) [15]. 2.6. Statistical analysis All the results were expressed as the Mean ± S.D. 3. Results and discussion The plant material was found to contain the required major phyto-compounds and other nutritive compounds desirable by
the pharmaceutical companies as well as in food production companies as supplements. Medicinal plants constitute the major constituents of most indigenous medicines and a large number of allopathic medical preparations enclose one or more components of plant origin [6]. In the current study, we screened for phytoconstituents present in A. precatorius and showed the presence of various classes of secondary metabolites such as alkaloids, flavonoids, phenols, tannins and steroids. Medicinal plants contain some organic compounds which provide definite physiological action on the human body and these bioactive substances include tannins, alkaloids, carbohydrates, terpenoids, steroids and flavonoids. Knowledge of the chemical constituents of plants is desirable because such information will be value for synthesis of complex chemical substances [16]. H-NMR-based analysis was sufficient to generate data from a sample within a relatively short time. H-NMR and C13 NMR spectra of A. precatorius revealed several signals as shown in Fig. 1. The bioactive compounds in the crude extract were individually characterized by their respective chemical shifts values as shown in the spectra (Fig. 1 A&B). Profiling of the bioactive principles of A. precatorius whole plant by H-NMR and C-13 NMR revealed the presence of different secondary metabolites. Despite the limitations for the analysis of crude extracts by NMR due to overlapping signals, we observed consistencies in our GC–MS fingerprints and NMR spectra, based on the detection of phyto-compounds and/or derivatives with similar pharmacological activities. Methanol extracts of A. precatorius leaves was subjected to GC–MS studies and the profile was shown in (Fig. 2). The results were occurred in 11 peaks which indicated the presence of 9 phytochemical constituents. On the comparison of the mass spectra of the constituents with the NIST library, the 9 compounds were characterized and identified (Table 1). Therefore, A. precatorius is recommended as a plant of phyto-pharmaceutical importance on account of the abundance level of major phyto-compounds that may be utilized by drug-designers following appropriate isolation and characterization procedures for the active principles [17]. Free radicals are known to play a definite role in a wide variety of pathological manifestations. Antioxidants fight against free radicals and protect us from various diseases. They exert their action either by scavenging the reactive oxygen species or protecting the antioxidant defence mechanisms [18,19]. In the current study, we evaluated the antioxidant capability of A. precatorius and leaves extract was demonstrated that significant free radical scavenging activity. On the other hand, we estimated the enzymatic antioxidants efficacy such as SOD, CAT and GPx and found that the A. precatorius leaves extracts shown the noteworthy antioxidant potentiality (Table 2). The electron donation ability of natural
Fig. 1. (A) C-13 NMR spectra of A. precatorius and (B) H-NMR spectrum of A. precatorius.
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
3
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
Fig. 2. The spectrum of bioactive compounds present in A. precatorius leaves methanolic extract.
Table 1 GC–MS spectral analysis of the extract of A. precatorius. S. No
Names of Compound
Molecular Formula
Molecular weight
Retention Time (RT) (Min)
Area (%)
1 2 3 4 5 6 7
Dimethyl Sulfoxide Octadecanal 3,7,11,15-Tetramethyl-2-Hexadecen-1-OL 9-OxononanoicAcid Squalene DI-N-Decylsulfone 2H-1-Benzopyran-6-OL, 3,4-Dihydro-2,5,7,8-Tetramethyl-2-(4,8,12Trimethy 1,2-BIS(Trimethylsilyl)Benzene 15,17-Dotriacontadiyne
C2H6OS C18H36O C20H40O C9H16O3 C30H50 C20H42O2S C31H52O3
78 268 296 172 410 346 472
11.112 18.330 18.750 21.811 25.342 27.193 27.763
6.117 12.407 18.439 4.556 5.393 2.465 20.266
C12H22Si2 C32H58
222 442
30.074 13.253
3.081 30.455
8 9
Table 2 Antioxidant capability of A. precatorius leaves methanolic extract. Concentration (lg/mL)
25 50 100
% Inhibition DPPH
ABTS
45.24 ± 1.25 59.39 ± 1.58 79.58 ± 2.11
39.58 ± 1.08 62.25 ± 2.01 85.65 ± 2.41
SOD (Units/mg protein)
Catalase (mmole of H2O2 consumed/min/mg protein)
GPx (mg of glutathione oxidized/min/mg protein)
21.25 ± 0.55 29.25 ± 0.64 38.25 ± 0.58
34.25 ± 0.29 41.21 ± 0.57 52.25 ± 0.61
221.20 ± 3.21 175.52 ± 2.89 128.25 ± 3.56
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
4
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
products can be measured by DPPH purple-colored solution bleaching [20]. The method is based on scavenging of DPPH through the addition of a radical species or antioxidant that decolorizes the DPPH solution. In the present study methanol extract showed significantly higher inhibition percentage and positively correlated with total phenolic content. The results of in vitro studies suggest that methanolic extract of A. precatorius may be useful in defence against free radicals, possibly be due to its antioxidant properties. To elevate the damaging effects of ROS, plants have evolved intracellular enzymatic antioxidants that include SOD, CAT and GPx. Plants are still a large source of natural antioxidants that might serve as leads for the development of novel drugs [21]. Super oxide radicals are inactivated by the enzyme SOD, the only enzyme known to use a free radical as a substrate. The radical scavenging activity of SOD is effective only when it is followed by increase in activity of CAT and other peroxidases. SOD generates H2O2 as a product which is in turn more toxic to the cells and requires CAT or peroxidases to scavenge. Thus a concomitant increase in CAT and or peroxidase is essential for the beneficial effect from increase SOD activity. CAT act in the micro body of cells, while GPx exists in the apoplast, chloroplast and cytosol [22]. It is understood that defensive mechanism against oxidative stress is primarily dependent upon orchestra synergism between exogenous and endogenous antioxidants. The exogenous antioxidants like Vitamin E and Vitamin C are recycled continuously by thiols like glutathione and dihydrolipoate. Thus, vitamin C and glutathione react cooperatively in vivo leading to greater protection against radical damage which could not be provided any single antioxidant [23]. Thus, A. precatorius contain all the antioxidative enzymes which can regulate the free radical activity and can reduce the generation of free radicals and may prevent cellular and tissue damage in human body. 4. Conclusion From the current study we identified an array of bioactive compounds present in A. precatorius and reported their antioxidant capability to effectively fight against free radicals. Therefore, A. precatorius is recommended as a plant of phyto-pharmaceutical importance on account of the abundance level of major
phyto-compounds that can be utilized by drug-designers following appropriate isolation and characterization procedures for the active principles. Acknowledgements The authors are thankful to the Indian Council of Medical Research (ICMR), Government of India for financial assistance. References [1] Tushar Dhanani, N.A. Sonal Shah, Satyanshu Kumar Gajbhiye, Arabian J. Chem. 10 (2017) 1193–1199. [2] G. Joana Gil-Chávez, J.A. Villa, J. Fernando Ayala-Zavala, et al., Compr. Rev. Food Sci. Food Saf. 12 (2013) 5–23. [3] Xu Dong-Ping, Ya Li, Xiao Meng, Tong Zhou, Yue Zhou, Jie Zheng, Jiao-Jiao Zhang, Hua-Bin Li, Int. J. Mol. Sci. 18 (2017) 96. [4] V.V. Sathibabu Uddandrao, P. Brahmanaidu, M. Balaji, G. Saravanan, Oxid. Antioxid. Med. Sci. 5 (2016) 79–86. [5] R. Szymanska, P. Pospíšil, J. Kruk, Oxid. Med. Cell. Longev. 2018 (2018) 2068370. [6] Uddandrao Sathibabu, G. Saravanan Brahmanaidu, Cardiovasc. Hematol. Agents Med. Chem. 15 (2017) 71–77. [7] M. Bhatia, N.A. Siddiqui, S. Gupta, Indo Am. J. Pharm. Res. 3 (2013) 3295–3315. [8] P. Brindha, B. Sasikala, K.K. Purushothaman, Bull. Med. Eth. Bot. Res. 3 (1981) 84–96. [9] R.C. Breton, W.F. Reynold, Nat. Prod. Rep. 30 (2013) 501–524. [10] B.P. Ezhilan, R. Neelamegam, Pharmacogn. Res. 4 (2012) 11–14. [11] W. Brand-Williams, M.E. Cuvelier, C. Berset, Lebens- Wiss. Technol. 28 (1995) 25–30. [12] R. Re, N. Pellegrini, A. Proteggente, M. Yang, C. Rice-Evans, Free Radic. Biol. Med. 26 (1999) 1231–1237. [13] A.K. Sinha, Anal. Biochem. 47 (1972) 389–394. [14] J.T. Rotruck, A.L. Pope, H.E. Ganther, A.B. Swanson, D.G. Hafeman, W.G. Hoekstra, Science. 179 (1973) 588–590. [15] K. Das, L. Samanta, G.B.N. Chainy, Ind. J. Biochem. Biophys. 37 (2000) 201–204. [16] V.V. Sathibabu Uddandrao, P. Brahmanaidu, P.R. Nivedha, S. Vadivukkarasi, G. Saravanan, Cardiovasc. Toxicol. 18 (2018) 199–205. [17] Philip Ilani, Nicholas Ajodo, Folashade Adewusi, et al., J. Coastal Life Med. 4 (2016) 395–402. [18] V.V. Sathibabu Uddandrao, P. Brahmanaidu, R. Ravindarnaik, et al., Eur. J. Nutr. (2018), https://doi.org/10.1007/s00394-018-1795-x. [19] V.V Sathibabu Parim G Saravanan Uddandrao Heart Fail Rev. (2018) 10.1007/ s10741-018-9749-1. [20] P.X. Nunes, S.F. Silva, R.J. Guedes, Venketeshwer Rao (Ed.), Global Approaches to Their Role in Nutrition and Health (2012), ISBN: 978-953-51-0203-8. [21] T. Xue, H. Hartikainen, V. Piironen, Plant Soil 237 (2001) 55–61. [22] D. Harman, Proc. Natl. Acad. Sci. 88 (1991) 5360–5363. [23] M. Gul, F.Z. Kutay, S. Temocin, Hanninen, Indian J. Exp. Biol. 38 (2000) 625– 634.
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
Contents lists available at ScienceDirect
Materials Today: Proceedings journal homepage: www.elsevier.com/locate/matpr
Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity K. Pavithra 1, V.V. Sathibabu Uddandrao 1, S. Mathavan, N. Gobeeswaran, S. Vadivukkarasi, Saravanan Ganapathy ⇑ Centre for Biological Sciences, Department of Biochemistry, K. S. Rangasamy College of Arts and Science (Autonomous), Tiruchengode, Namakkal District, Tamilnadu 637215, India
a r t i c l e
i n f o
Article history: Received 7 April 2019 Received in revised form 9 May 2019 Accepted 13 May 2019 Available online xxxx Keywords: Abrus precatorius Bioactive compounds Antioxidants Medicinal plants Natural products
a b s t r a c t The current study was designed to develop fingerprint of the different bioactive compounds of Abrus precatorius and evaluation of its antioxidant activity. C-13 and H-NMR analysis of A. precatorius elucidated the key bioactive compounds in the leaves that were consistent with the classes of bioactive compounds detected by GC–MS analysis which revealed the presence of 9 major compounds. A. precatorius shown significant free radical scavenging activities and contain antioxidative enzymes which can regulate the free radical activity. In conclusion, the present study identified an array of bioactive compounds present in A. precatorius and reported their ethno-botanical uses against free radicals. Ó 2019 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Emerging Materials and Modeling.
1. Introduction Plants are an imperative source of bioactive molecules for drug discovery. Isolated bioactive molecules serve as starting materials for laboratory synthesis of drugs as well as a model for the production of biologically active compounds [1]. The search for natural bioactive compounds with prospective for the treatment and prevention of human diseases and to meet other needs is presently a key topic in many laboratories and industries. These compounds competently interact with DNA, proteins and other biological molecules to produce a desired outcome, which could be exploited for designing natural products-derived beneficial agents [2]. The study of medicinal plants starts with the pre-extraction and the extraction procedures, which is an important step in the processing of the bioactive constituents from plant materials. Considering their significant health effects, the proficient extraction methods of natural antioxidants, suitable assessment of antioxidant activity as well as their major resources from medicinal and food plants are depict enormous attention in food science and nutrition [3]. Plant-derived antioxidants are a large group of natural products with reducing or radical-scavenging capacity [4]. Due to their potent defensive, as well as medicinal actions, these compounds ⇑ Corresponding author. 1
E-mail address: [email protected] (S. Ganapathy). Both authors contributed equally to this work.
receive a great deal of attention by not only scientists but also pharmacologists and physicians [5]. In present time, medicinal plants as rich source of natural bioactive components are given precedence to study their antioxidant activity and discover their utilization in treatment of diabetes mellitus, dyslipidemia and cardiovascular diseases [6]. Abrus precatorius (Fabaceae) which is an ornamental, twining and woody vine originally native to India is now commonly found throughout the subtropical and tropical parts of the world [7]. In view of the above literature, we made an attempt to extract and identify the bioactive factors from A. precatorius by GC–MS and NMR and evaluated its antioxidant capability through in vitro studies.
2. Materials and methods 2.1. Sample preparation The fresh leaves of A. precatorius were collected from Tamilnadu, India and the plant was authenticated by Botanical Survey of India (BSI/SRC/5/23/2017/Tech/2874). The leaves were shade dried at 7 days and pulverized to powder in a mechanical grinder and extracted by continuous extraction using soxhlet apparatus with methanol for 72 h. The extract was collected and solvents were evaporated and dried in hot air oven at 50 °C–100 °C. The dried powder was collected and stored for further analysis.
https://doi.org/10.1016/j.matpr.2019.05.417 2214-7853/Ó 2019 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Emerging Materials and Modeling.
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
2
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
2.2. Preliminary phytochemical analysis The A. precatorius extract was subjected to qualitative examination for different phytoconstituents as per standard procedures [8]. 2.3. Nuclear magnetic resonance (NMR) analysis Hydrogen-NMR spectra of the extracts was performed using Bruker Biospin Avance 400-MHz NMR spectrophotometer with a 5 mm broad inverse probe head, equipped with shielded z-gradient accessories. Hydrogen-NMR spectral analysis of the extract was carried out using one-pulse sequence by dissolving the extract in 500 lL of deuterated methanol in 5 mm NMR tubes. A re-usable, sealed capillary tube containing 30 lL of 0.375% of trisodium phosphate in deuterium oxide was inserted into the NMR tube before recording the spectra. Tri-sodium phosphate served as chemical shift reference as well as internal standard for quantitative estimation. C-13 NMR heteronuclear single quantum correction was carried out using the Brucker’s standard pulse library [9]. 2.4. Gas chromatography-mass spectrometry (GC–MS) analysis The Clarus 680 GC was used in the analysis employed a fused silica column, packed with Elite-5MS (5% biphenyl 95% dimethylpolysiloxane, 30 m X 0.25 mm ID X 250 lm df) and the components were separated using Helium as carrier gas at a constant flow of 1 mL/min. The spectrums of the components were compared with the database of spectrum of known components stored in the GC–MS National Institute Standard and Technology (2008) library [10]. 2.5. Determination of in vitro antioxidant activity We assessed the antioxidant activity of A. precatorius extract by 2, 20 -diphenyl-1- picrylhydrazyl (DPPH) radical scavenging assay [11], 2,20 -azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging activity [12], activities of catalase (CAT) [13], glutathione peroxidase (GPx) [14] and superoxide dismutase (SOD) [15]. 2.6. Statistical analysis All the results were expressed as the Mean ± S.D. 3. Results and discussion The plant material was found to contain the required major phyto-compounds and other nutritive compounds desirable by
the pharmaceutical companies as well as in food production companies as supplements. Medicinal plants constitute the major constituents of most indigenous medicines and a large number of allopathic medical preparations enclose one or more components of plant origin [6]. In the current study, we screened for phytoconstituents present in A. precatorius and showed the presence of various classes of secondary metabolites such as alkaloids, flavonoids, phenols, tannins and steroids. Medicinal plants contain some organic compounds which provide definite physiological action on the human body and these bioactive substances include tannins, alkaloids, carbohydrates, terpenoids, steroids and flavonoids. Knowledge of the chemical constituents of plants is desirable because such information will be value for synthesis of complex chemical substances [16]. H-NMR-based analysis was sufficient to generate data from a sample within a relatively short time. H-NMR and C13 NMR spectra of A. precatorius revealed several signals as shown in Fig. 1. The bioactive compounds in the crude extract were individually characterized by their respective chemical shifts values as shown in the spectra (Fig. 1 A&B). Profiling of the bioactive principles of A. precatorius whole plant by H-NMR and C-13 NMR revealed the presence of different secondary metabolites. Despite the limitations for the analysis of crude extracts by NMR due to overlapping signals, we observed consistencies in our GC–MS fingerprints and NMR spectra, based on the detection of phyto-compounds and/or derivatives with similar pharmacological activities. Methanol extracts of A. precatorius leaves was subjected to GC–MS studies and the profile was shown in (Fig. 2). The results were occurred in 11 peaks which indicated the presence of 9 phytochemical constituents. On the comparison of the mass spectra of the constituents with the NIST library, the 9 compounds were characterized and identified (Table 1). Therefore, A. precatorius is recommended as a plant of phyto-pharmaceutical importance on account of the abundance level of major phyto-compounds that may be utilized by drug-designers following appropriate isolation and characterization procedures for the active principles [17]. Free radicals are known to play a definite role in a wide variety of pathological manifestations. Antioxidants fight against free radicals and protect us from various diseases. They exert their action either by scavenging the reactive oxygen species or protecting the antioxidant defence mechanisms [18,19]. In the current study, we evaluated the antioxidant capability of A. precatorius and leaves extract was demonstrated that significant free radical scavenging activity. On the other hand, we estimated the enzymatic antioxidants efficacy such as SOD, CAT and GPx and found that the A. precatorius leaves extracts shown the noteworthy antioxidant potentiality (Table 2). The electron donation ability of natural
Fig. 1. (A) C-13 NMR spectra of A. precatorius and (B) H-NMR spectrum of A. precatorius.
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
3
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
Fig. 2. The spectrum of bioactive compounds present in A. precatorius leaves methanolic extract.
Table 1 GC–MS spectral analysis of the extract of A. precatorius. S. No
Names of Compound
Molecular Formula
Molecular weight
Retention Time (RT) (Min)
Area (%)
1 2 3 4 5 6 7
Dimethyl Sulfoxide Octadecanal 3,7,11,15-Tetramethyl-2-Hexadecen-1-OL 9-OxononanoicAcid Squalene DI-N-Decylsulfone 2H-1-Benzopyran-6-OL, 3,4-Dihydro-2,5,7,8-Tetramethyl-2-(4,8,12Trimethy 1,2-BIS(Trimethylsilyl)Benzene 15,17-Dotriacontadiyne
C2H6OS C18H36O C20H40O C9H16O3 C30H50 C20H42O2S C31H52O3
78 268 296 172 410 346 472
11.112 18.330 18.750 21.811 25.342 27.193 27.763
6.117 12.407 18.439 4.556 5.393 2.465 20.266
C12H22Si2 C32H58
222 442
30.074 13.253
3.081 30.455
8 9
Table 2 Antioxidant capability of A. precatorius leaves methanolic extract. Concentration (lg/mL)
25 50 100
% Inhibition DPPH
ABTS
45.24 ± 1.25 59.39 ± 1.58 79.58 ± 2.11
39.58 ± 1.08 62.25 ± 2.01 85.65 ± 2.41
SOD (Units/mg protein)
Catalase (mmole of H2O2 consumed/min/mg protein)
GPx (mg of glutathione oxidized/min/mg protein)
21.25 ± 0.55 29.25 ± 0.64 38.25 ± 0.58
34.25 ± 0.29 41.21 ± 0.57 52.25 ± 0.61
221.20 ± 3.21 175.52 ± 2.89 128.25 ± 3.56
Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417
4
K. Pavithra et al. / Materials Today: Proceedings xxx (xxxx) xxx
products can be measured by DPPH purple-colored solution bleaching [20]. The method is based on scavenging of DPPH through the addition of a radical species or antioxidant that decolorizes the DPPH solution. In the present study methanol extract showed significantly higher inhibition percentage and positively correlated with total phenolic content. The results of in vitro studies suggest that methanolic extract of A. precatorius may be useful in defence against free radicals, possibly be due to its antioxidant properties. To elevate the damaging effects of ROS, plants have evolved intracellular enzymatic antioxidants that include SOD, CAT and GPx. Plants are still a large source of natural antioxidants that might serve as leads for the development of novel drugs [21]. Super oxide radicals are inactivated by the enzyme SOD, the only enzyme known to use a free radical as a substrate. The radical scavenging activity of SOD is effective only when it is followed by increase in activity of CAT and other peroxidases. SOD generates H2O2 as a product which is in turn more toxic to the cells and requires CAT or peroxidases to scavenge. Thus a concomitant increase in CAT and or peroxidase is essential for the beneficial effect from increase SOD activity. CAT act in the micro body of cells, while GPx exists in the apoplast, chloroplast and cytosol [22]. It is understood that defensive mechanism against oxidative stress is primarily dependent upon orchestra synergism between exogenous and endogenous antioxidants. The exogenous antioxidants like Vitamin E and Vitamin C are recycled continuously by thiols like glutathione and dihydrolipoate. Thus, vitamin C and glutathione react cooperatively in vivo leading to greater protection against radical damage which could not be provided any single antioxidant [23]. Thus, A. precatorius contain all the antioxidative enzymes which can regulate the free radical activity and can reduce the generation of free radicals and may prevent cellular and tissue damage in human body. 4. Conclusion From the current study we identified an array of bioactive compounds present in A. precatorius and reported their antioxidant capability to effectively fight against free radicals. Therefore, A. precatorius is recommended as a plant of phyto-pharmaceutical importance on account of the abundance level of major
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Please cite this article as: K. Pavithra, V. V. S. Uddandrao, S. Mathavan et al., Identification of bioactive factors from Abrus precatorius by GC–MS, NMR and evaluation of its antioxidant activity, Materials Today: Proceedings, https://doi.org/10.1016/j.matpr.2019.05.417