Author(s): N Senthilkumar* and R Sumathi
ABSTRACT
Pterocarpus santalinus (Red Sanders) a highly impressive indigenous deciduous threatened tree species renowned for its characteristic timber of exquisite colour reported to have enormous biological activities and potential health benefits along with its extensive use in furniture. Considering its various health benefits, the present study focused on evaluation of phytochemical constituents of the leaf, bark and heartwood of P. santalinus responsible for its anti-inflammatory activity. Phytocompounds were quantified high in heartwood samples collected from natural forests than plantations. Alkaloids and terpenoids were present in both acetone and methanol extracts of leaf, bark and heartwood samples accounts for potential medicinal uses especially its anti- inflammatory activity. GC/ MS/MS analysis revealed that the compounds elicited at various retention times found to have various biological properties such as antimicrobial, antioxidant, anti-inflammatory, anti-diabetic and anti-malarial activities. Molecular docking study was aimed to predict the affinity of the compounds namely Viminalol, Protopine, Lupanol, Lup-20(29)-en-3-one and Lup-20(29)-en-3-ol, (3.beta.)- (CAS) of red sanders to bind with anti-inflammatory targeted proteins viz. COX-1, COX-2, NR3C1, S100s, IL1F5 and SPINK5 thereby reduced their functionality towards inflammation. The results proved that red sanders found to have the potential as an anti-inflammatory agent and it can be used in the development of health care products.
The natural plant based remedies are as old as humankind which was evidenced through pre-historic sites and written records and plants are the source of first medicines. Plant based natural medicines are the most widely used medicines for both acute and chronic health issues even with the huge availability of antibiotics and enormous synthetic medicine [1-3]. Over the past 100 years, the development and bulk manufacture of chemically synthesized medicines have revolutionized health care in most parts of the word. In developing countries large population still rely on traditional practitioners and herbal medicines for their primary health care. During the past two decades, use of traditional medicine is not only limited to developing countries, public interest in natural therapies has increased greatly in industrialized countries also with expanding use of ethno botanicals [4]. The growing demand towards plant based natural products has renewed attention in the production of herbal health care formulations, herbal-based cosmetic products, and herbal nutritional supplements. Hence, there is a demand to promote natural products to save the human lives. Pullaiah et al. reported that Pterocarpus santalinus is one such tree species in which the bioactive compounds present in it accounts for its potential health benefits [5].
Pterocarpus santalinus L.f. belongs to the family Fabaceae, popularly known as Red Sanders is one of the threatened plants of India. It is an endemic and endangered timber tree species confined to Southern parts of Eastern Ghats. India is one of the countries which has the favourable soil and climatic conditions to grow red sanders. The natural distribution of red sanders occurs almost exclusively in the south eastern parts (Chittoor, Cuddapah and Nellore districts of Seshachalam hill ranges, Kurnool, Nellore and Prakasam) of Andhra Pradesh in the tropical dry deciduous forests and northern parts (North Arcot, and Chengalpet districts) of Tamil Nadu [6]. Red sanders is highly impressive indigenous herb, renowned for its characteristic timber of exquisite colour, beauty and exceptional technical qualities. It is a highly valued tree species for its diverse utility in India and abroad.
Its valuable wavy grained timber is traded as padauk, the wood is also used in traditional and folklore medicines and the red dye obtained from the wood is used as colouring agent for textile, medicine and food. Hence, the natural sources of red sanders were exploited indiscriminately and already included in the endangered category of IUCN red list [7]. Along with its extensive use in furniture, enormous biological activities and potential health benefits of P. santalinus have been reported, including antioxidant, anti-diabetic, antimicrobial, anticancer, anti-inflammatory, anti-tumour and protective effects on the liver, gastric mucosa, nervous system, prickly heat, skin diseases, snake bite, Jaundice and various other ailments were attributed to the bioactive compounds present in it [6].
Significant bioactive compounds identified from P. santalinus with promising medicinal uses reported to have more subtle effects on cellular mechanism which reduces the effects of disease instead of preventing deficiency diseases. Stem, bark and leaf extracts of P.santalinus showed maximum antimicrobial activity [8, 9]. Considering its various health benefits, the present study focused on the evaluation of phytochemical constituents of the leaf, bark and heartwood of P. santalinus responsible for its anti- inflammatory activity.
Inflammatory activity is caused by wound and infection in the tissues because of which the immune cells will release nitric oxide (NO), prostaglandin (PG), interleukin 6 (IL-6), and tumor necrosis factor-α (TNF-α) in human body [10,11]. Glucocorticoid receptor, NR3C1 is a protein that is expressed constitutively and ubiquitously throughout the body and transcriptional regulator of anti-inflammatory molecules which upon ligand binding enters the nucleus and control diverse biological processes including development, metabolism and inflammation [12]. Activated glucocorticoid receptor up-regulates the expression of anti- inflammatory proteins in the nucleus [13]. Prostaglandin is a major mediator of inflammation process and its excessive production via up-regulation of the COX-2 activity leads to inflammatory mediated diseases including cancer Alzheimer’s diseases and acute renal failure [14-18]. Cyclooxygenase (COX) is the first stimulus enzyme involved in the prostaglandin synthetic pathway. COX-1 also causes inflammation though housekeeping enzyme due to its constitutive role in human physiology [19-22].
Inhibition of COX-2 using natural compounds will be a target for inflammation treatment with lesser side effects [23-27]. Selective COX-2 inhibitors using the bioactive compounds isolated from natural resources is preferable to reduce the undesirable side effects of the long term usage of synthetic drugs [28]. Wang et al. reported that the active compound eburicoic acid isolated from Laetiporus sulphureus showed significant anti-inflammatory activity. Platycodi radix root is widely consumed for ocular inflammation due to high content of saponin [29-32].
Plant based bioactives can also be predicted through molecular docking, an in silico approach other than biochemical and chromatographic profiling. The primary purpose of this method is to predict the affinity of a drug candidate (ligand) to bind with the protein and form the most stable complex thereby reduce or block its activity which is responsible for various causative illness. Molecular docking is one of the vital computational methodology and a valuable tool used widely by researchers to underscore the molecular interactions of ligand molecules for drug discovery and development trajectory [33,34]. Hence, in the present study bioactive compounds present in the leaf, bark and heartwood of red sanders having significant biological activities were identified through phytochemical screening, chromatographic analysis and molecular docking studies to confirm the anti-inflammatory activity of red sanders.
Extensive surveys have been made in Southern states of India viz. Tamil Nadu, Andhra Pradesh and Karnataka for collection of leaf, bark and heartwood samples of P. santalinus. Samples were collected from plantations in Tanjore, Karaikudi, Dharmapuri and Thiruvannamalai districts of Tamil Nadu; natural strands in Rajampet, Tirupati and YSR districts of Andhra Pradesh and plantations Bangalore, and Kolar districts of Karnataka. The plant samples were washed thoroughly with tap water and shade dried for a week at room temperature (24 ± 2°C). The dried samples were powdered to get coarse granules using an electric blender and stored in an airtight container for further analysis.
The powdered plant samples (leaf, bark and heartwood) of P. santalinus (50 g) were extracted with 350 ml of acetone and methanol using Soxhlet apparatus at 65-80 °C for 8-10 h to extract the phytocompounds. The solvents of the respective extracts were reduced in rotary vacuum evaporator until complete removal of solvents and the extracts obtained were stored at 4 °C until further use. The stored plant extracts were then dissolved in respective solvents while experimentation to get the working solution of 1 mg/1 mL which has been subjected to qualitative and quantitative analysis.
Qualitative phytochemical screening of the methanol and acetone extracts of leaf, bark and heartwood samples of P. santalinus collected from selected morphologically superior trees was carried out to identify the presence of various secondary metabolites such as Alkaloids, Flavonoids, Tannins, Saponins, Steroids, Phenols, Glycosides, Protein, Carbohydrate and Terpenoids using standard methods [35].
The total alkaloid content present in the leaf, bark and heartwood of P. santalinus was estimated using method described by Karawya et al. Alkaloid present in the sample reacted with bromocresol green and produced yellow colour complex and measured spectrophotometrically at 415 nm [36].
The flavonoid content of bark and heartwood extracts of P. santalinus was estimated by the method of Harborne [35]. Flavonoids present in the sample react with vanillin reagent in acidic medium and produced yellow colour which was measured spectrophotometrically.
The total phenolic compound present in the bark and heartwood of P. santalinus was estimated Folin-Ciocalteu reagent by the method described by Malick and Singh [37]. Phenols reacted with phosphomolybdic acid in Folin-Ciocalteu reagent in alkaline medium and produced blue coloured complex called molybdenum blue.
The saponin content present in the leaf, bark and heartwood samples of P. santalinus was estimated by the method of Makkar et al. The saponin was quantified based on the colorimetric reaction of vanillin-sulphuric acid with slight modifications [38].
Tannin like compounds reduce phosphotungstate to molybdic acid in alkaline solution to produce a blue colour complex and the colour intensity is proportional to the concentration of tannin and measured at 700 nm [39].
Steroids reacted with ferric chloride in the presence of concentrated sulphuric acid gave pink colour. The intensity of colour developed is directly proportional to the amount of steroids present and it was read at 540 nm in a calorimeter [40].
The GC-MS/MS analysis was performed using Varian 3800 with Mass spectrum 4000 GC-MS/MS system equipped with a Fused silica capillary column of size 15m x 0.2 mm ID x 1μm linked to an EI detector. Helium gas (99.99% purity) was used as a carrier gas at a constant flow rate of 1ml/min and the sample injected was 1μl; the instrument was set to an initial temperature of 110 ºC, and maintained at this temperature for 2 min. At the end of this period the oven temperature was rose up to 280 ºC, at the rate of an increase of 5 ºC/min, and maintained for 9 min. Injection port temperature was ensured as 250 ºC. The ionization voltage was 70eV. The samples were injected in split mode as 10:1. Mass spectral scan range was set at 45-450 (m/z). Using computer searches on a NIST Ver.2.1 MS data library and comparing the spectrum obtained through GC-MS/MS compounds present in the plants sample were identified.
The active components in the extracts were identified by comparing their retention indices and mass spectra patterns with the spectrum of known components stored in the computer library and also with published literatures. Interpretation on mass-spectrum GC-MS/MS was conducted to match the identified components from the plant material using the database of National Institute of Standard and Technology (NIST), Wiley, Mainlib, Replib and Tutorial library sources having more than 62,000 patterns. The name, molecular weight and structure of the components of the test materials were ascertained.
The extract yield was comparatively high in methanol than acetone extracts in all three plant samples viz. leaf, bark and heartwood of red sanders. In leaf samples methanol extract yield was high in samples collected from Thiruvannamalai (14.19 %) district of Tamil Nadu followed by YSR district of Andhra Pradesh (11.12 %). In case of bark samples the yield was high in methanol extract of samples collected from Rajampet, Andhra Pradesh (14.45 %) followed by Thiruvannamalai (14.19 %) district of Tamil, Nadu (12.64%). In case of heartwood the yield was high in methanol extract of samples collected from YSR district of Andhra Pradesh (16.15 %) followed by Rajampet, Andhra Pradesh (12.34 %). Over all the extract yield was high in heartwood, followed by bark and leaf the least (Figure. 1).
Phytochemical screening of methanol and acetone extracts of leaf, bark and heartwood of P. santalinus showed the presence of alkaloids, tannins, saponins, phenols and terpenoids. Methanol elicited more phytocompounds than acetone invariably in leaf, bark and heartwood. The samples collected from Karaikudi, Dharmapuri and Thiruvannamalai screened for maximum compounds of 6. Among all plant samples heartwood elicited more compounds followed by bark and leaf the least. Dharmapuri sources found to have tannins in both acetone and methanol extracts of leaf and bark. Alkaloids, terpenoids were present in both acetone and methanol extracts of leaf, bark and heartwood samples of all sources (Table 2).
Quantitative analysis of phyto-chemicals in leaf, bark and heartwood extracts of P. santalinus showed that among all phytochemicals phenols was quantified high, followed by flavonoids and alkaloids and acetone quantified more than methanol extract samples. Heartwood contains more of phenols, alkaloids compared to bark and leaves. Bark contains more saponin and tannin than leaf and heartwood. Comparatively Tirupati (Andhra Pradesh) samples contain high quantity of alkaloid (101.69 mg/g), flavonoid (135.13 mg/g), tannin (99.33 mg/g) and saponin (118.78 mg/g) than other location samples. Steroids (107.14 mg/g) content were quantified high in YSR district, Andhra Pradesh samples. Phenol (227.47 mg/g) content were quantified high in Dharmapuri samples
GC/MS/MS analysis of the acetone and methanol extract of leaf, bark and heartwood (Fig 2, 3 & 4) showed the presence of various bioactive compounds with medicinal properties by relating their peak retention time, peak area (%), height (%) and mass spectral fragmentation profile with the known compounds described by the National Institute of Standards and Technology (NIST) and Wiley libraries. The GC/MS/MS chromatogram of methanol extracts of leaf, bark and heartwood samples of P. santalinus recorded more bioactive compounds in heartwood followed by bark and leaf. Results revealed that 4, 10 and 24 compounds were elicited in leaf, bark and heartwood methanol extract of P. santalinus respectively (Table 3, 4 & 5). The phytocompounds 2-Decyl-1H-quinolin-4- one separated at RT 27.649 was quantified high (34.748 %) followed by Gomezine (27. 973 5) in leaf. In bark lupanol was recorded high (22.743 %) followed by 8,13-epoxy-labadan-1,6,7,9-tetraol-11-one, 7-o-acetate(ester) (12.025 %). Maximum of 24 compounds were found in the heartwood of red sanders collected from Dharmapuri (Table 5). In heartwood, 9.beta.-Acetoxy-4-hydroxy-3,4,8- trimethyl-5.alpha.-H-tricyclo[6.3.1.0(1,5)] dodecane was recorded in high quantity (16.992 %). The compounds Viminalol, Protopine, Lup-20(29)-en-3-one, Lup-20(29)-en-3-ol, (3.beta.)- (CAS) identified in heartwood and lupanol in bark samples of red sanders were reported to have anti-inflammatory activity. Hence these compounds were subjected to molecular docking studies for further confirmation through in silico cost effective study.
Figure 4: GC/MS/MS Chromatogram of Methanol Extract Red Sanders Heartwood
In the present study, molecular interaction of the bioactive compounds viz. Viminalol, Protopine, Lupanol, Lup-20(29)-en-3-one, Lup-20(29)-en-3-ol, (3.beta.)- (CAS) having anti-inflammatory activity with 6 anti-inflammatory target proteins namely COX-1, COX-2, NR3C1, S100s, IL1F5 and SPINK5 was carried out on Dell Optiplex 380 Intel (R) Core (TM) i-2400 CPU @ 3.10 GHz processor. Molecular docking studies of the active compounds of heartwood extract of red sanders identified though GC/MS/MS chromatographic technique having promising biological activities revealed that few compounds showed significant docking score for anti-inflammatory activity. All the bioactive compounds used for molecular docking against NR3C1 (1NHZ), COX-2 (5FIA) and COX-1(6Y3C) showed significant docking scores (Fig 5). Especially Lup-20(29)-en-3-ol, (3.beta.)- (CAS) showed high docking score and can be used as drug against inflammation against NR3C1(1NHZ) and COX-2 (5FIA) efficiently.
During the past few decades interest in natural therapies has increased greatly worldwide with expanding use of ethnobotanicals [4]. Array of efficient nutrient benefits can be extracted in whole plant than the few isolated bioactive constituents from synthetic components. Phytochemicals contained in plants and tree species act as a defence system to combat diseases and benefit consumer health. The thirst towards natural therapies drive us to study the bioprospecting potential of red sanders. Among the three explants of P. santalinus used for the study, the extract yield was comparatively high in heartwood followed by bark and leaf. Invariably methanol extracted more of phytochemicals than acetone in all samples.
Senguttuvan et al. reported that methanol extract given higher percentage of yield than petroleum ether, chloroform, ethyl acetate, ethanol and water extracts of leaf and root of medicinal herb, Hypochaeris radicata [41]. Comparatively methanol extracted high variety of plant constituents due to its high polarity than other solvents [42]. Truong Dieu-Hien et al. reported that the extraction yield was high (33.2%) when the plant samples were extracted with methanol than ethanol, acetone, chloroform, and dichloromethane showing that the extraction efficiency was high in highly polar solvent [43]. Among the samples collected from various locations situated in Tamil Nadu, Andhra Pradesh and Karnatak the extract yield was recorded high in heartwood sample collected from YSR Districts (16.15 %) of Andhra Pradesh. Plant samples of red sanders collected from natural forest located in all three districts of Andhra Pradesh invariably gave high yield than samples collected from the plantations.
Phytochemical constituents in the medicinal plants possessing various potential biological activities were extensively used in the human treatment, veterinary, agriculture, scientific research and countless other areas [44-46]. Plants and its parts viz. leaf, stem, bark, fruit, root, twig, wood and sap have been used in traditional and as folk medicines by locals to cure several diseases which includes cough, fever, asthma, diarrhoea, indigestion, and skin diseases [47]. Plants reported to have low toxic bioactive organic compounds like alkaloids, tannins, flavonoids, terpenoids, saponins and phenolic compounds with therapeutic potential [48]. Phytochemical screening of methanol and acetone extracts of leaf, bark and heartwood of P. santalinus was carried out. It was observed that methanol elicited more of phytocompounds than acetone and among the plant samples heartwood elicited more compounds followed by bark and leaf.
Dharmapuri sources found to have tannins in both acetone and methanol extracts of leaf and bark. Alkaloids, terpenoids were present in acetone and methanol extracts of leaf, bark and heartwood samples of all sources. Arunakumara et al. confirmed the presence of various components, such as carbohydrates, steroids, anthocyanins, saponins, tannins in P. santalinus. The presence of flavonoids, glycosides, and phenols were reported in the bark of P. santalinus, alkaloids, flavonoids, tannins, phenols, steroids, saponins, glycosides, carbohydrates and terphenoids and carbohydrates, flavonoids, terpenoids, phenolic compounds, alkaloids, saponins, tannins, and glycosides were reported in the heartwood of red sanders [49-52].
A variety of plant ingredients with diverse structures are capable of promoting health benefits. Quantitative analysis of phyto- chemicals in leaf, bark and heartwood extracts of P. santalinus showed that among all phytochemicals phenols was quantified high, followed by flavonoids and alkaloids and acetone quantified more than methanol extract samples. Heartwood contains more of phenols, alkaloids compared to bark and leaves. Bark contains more saponin and tannin than leaf and heartwood. Compared to heartwood, bark have large amount of Tannins, phenols, alkaloids and flavonoids. Tannin had more application in industries and biological properties as antioxidants, antiseptics, anticarcinogenic and anti-inflammatory activity. Saponin has been responsible for some biological activities such that insecticidal activity, anthelmintic activity, pesticidal activity, antibacterial, antifungal and antiviral activity [53,54]. In pharmacology, phenol content in plants was mainly targets to treat different diseases [55].
Prolong usage of the drugs against any diseases have been associated with serious and sometimes life threatening side effects which emphasized the research to find out plant based alternative therapeutic regimen with comparative efficacy but with fewer side effects. The natural products with medicinal properties have been used to treat all sorts of inflammatory conditions [56,57]. Keeping this in view chromatographic identification of bioactives with significant biological properties pushed towards GC/MS/ MS analysis of leaf, bark and heartwood of red sanders. The compounds elicited at various retention time found to have various biological properties such as antimicrobial, antioxidant, anti-inflammatory, anti-diabetic and anti-malarial activities. The phytocompound Gomezine, a monoterpenoid indole alkaloid elicited at retention time of 30.705 reported to have cytotoxicity and analgesic properties [58,59]. Similarly an alkaloid berberine found in Berberis species, tryptanthrin in Isatis species reported to have anti-inflammatory activity and alkaloid in Cissampelos pareira accounts for its immunosuppressive and antioxidant activities [60-62].
The bioactive compound lupanol, natural pentacyclic triterpenoid identified in the bark of red sanders reported to have anti- inflammatory, anti-arthritic, anti-mutagenic and anti-malarial activity. The compound Viminalol a pentacyclic triterpenoid, Lup-20(29)-en-3-one, Lup-20(29)-en-3-ol, (3.beta.)- (CAS) and Protopine a isoquinoline alkaloid identified though GC/MS/MS analysis reported to have anti-thrombotic, anti-inflammatory, anti- spasmodic, neuroprotective, anti-microbial, anti-bacterial, anti- viral, anti-fungal and anti-parasitic activities. Viminalol isolated from the methanol extract of Artocarpus communis roots reported to have antimicrobial activity and antinociceptive and anti- inflammatory [63,64]. Protopine has been found to have diverse biological activities such as anti-inflammation, anti-microbial, anti-angiogenic and anti-tumours activities [65].
Among the phytocompounds identified through GC/M/MS analysis having various promising biological activities five of the compounds, Viminalol, Protopine, lupanol, Lup-20(29)-en- 3-one and Lup-20(29)-en-3-ol, (3.beta.)- (CAS) reported to have significant anti-inflammatory activity were selected for molecular docking studies to validate the compounds anti-inflammatory activity against the anti-inflammatory targeted proteins viz. COX- 1, COX-2, NR3C1, S100s, IL1F5 and SPINK5 using Dell Optiplex 380 Intel (R) Core (TM) i-2400 CPU @ 3.10 GHz processor suggested that all legand compounds against NR3C1 (1NHZ), COX-2 (5FIA) and COX-1(6Y3C) showed significant docking scores. Docking study to validate anti-inflammatory activity of the bioactives cemented the acanthoic acid a novel pimarane diterpene isolated from Acanthopanax kiusianus through its potential to suppress the COX-1 and COX-2 expressions thereby inhibiting prostaglandin synthesis [66]. COX-2 inhibitory activity of berberine extracted from Berberis species proved that it can be a therapeutic option in treating inflammation [67- 69]. In the present study, Lup-20(29)-en-3-ol, (3.beta.)- (CAS) in leaf, bark and heartwood of red sanders with high docking score proved to use as anti-inflammatory drug against COX-2 (5FIA) efficiently.
Pterocarpus santalinus is a native tree species of India endemic to Rayalaseema region of Andhra Pradesh for high value timber and furniture. It also contains many potent phytochemicals for use in pharmaceuticals and cosmetics industries in addition to its timber and industrial uses. The presence of alkaloids and terpenoids in leaf, bark and heartwood of P. santalinus projected it as a promising source of naturally occurring antioxidants especially accounts for its anti-inflammatory activity. This study affirms that the presence of phytocompounds with significant biological activities accounts for the effectiveness of red sanders in treating inflammation. Molecular docking studies highlighted the probable mechanism by which red sanders is expected to elicit anti-inflammatory effects which assures its natural therapeutic use in healing inflammation without side effects.
We acknowledge National Biodiversity Authority (NBA), Chennai for the funding support through a project no. Tech./ Genl/22/149/17/18-19/4295 dt 17.03.2020. We thank the Director, IFGTB for the facilities provided to conduct the study.
1.Hussain MS, Fareed S, Ali M (2010) Hygrophila auriculata (K. Schum) Heine: Ethnobotany, phytochemistry and pharmacology. Asian J. Tradit Med 5: 122-131.
2.Hussain MS, Fareed S, Ali M (2011) Preliminary phytochemical and pharmacognostical screening of the ayurvedic drug Hygrophila auriculata (K. Schum) Heine. Pharmacog J 3: 28-40.
3.Hussain MS, Fareed S, Ansari S, Iffat Z, Rahman A, et al. (2012) Current approaches toward production of secondary plant metabolites. J Pharm. Bioallied Sci 4: 10-20.
4.Wachtel Galo S, Benzie IFF (2011) Herbal Medicine: An introduction in its history, usage, regulation, current trends and research needs. In (Eds. Iris F. F. Benzie and SissiWachtel- Galo) Herbal Medicine: Biomolecular and Clinical Aspects, Second Edition. CRS Press 1-9.
5.Pullaiah T, Balasubramanya S, Anuradha M (2019) Red Sanders: Silviculture and Conservation. Springer, Singapore 210.
6.GOI (Government of India) (2014) Government of India. Ministry of Environment of Forests Publications. Red Sanders. Available at http://moef.nic.in/downloads/public- information /Chap-8-new.pdf.
7.IUCN (2014) IUCN Red list of threatened species. https:// www.iucn.org/resources/conservation-tool/iucn-red-list- threatened-species.
8.Arunkumara KKIU, Walpola BC, Subasinghe S, Yoon MH (2011) Pterocarpus santalinus Linn. f. (Rathhandun): a review of its botany, uses, phytochemistry and pharmacology, J. Korean Soc. Appl. Biol. Chem 54: 495-500.
9.Arunkumar AN, Joshi G (2014) Pterocarpus santalinus (red sanders) an endemic, endangered tree of India: current status. improvement and the future J. Trop for Environ 4: 1-10.
10.Zhang QL, Zhang J, Xia PF, Peng XJ, Li HL, et al. (2019) antiinflammatory activities of gentiopicroside against iNOS and COX-2 targets Chinese Herbal Medicines 11: 108-112.
11.Yatam S, Gundla R, Jadav SS, reddy Pedavenkatagari N, Chimakurthy J, et al. (2018) Focused library design and synthesis of 2-mercapto benzothiazole linked 1, 2, 4-oxadiazoles as COX2/5-LOX inhibitors. Journal of Molecular Structure 1159: 193-204.
12.Evans RM, Mangelsdorf DJ (2014) nuclear receptors, RXR, and the big bang. Cell 157: 255-266.
13.Rhen Turk, Cidlowski, John A (2005) Antiinflammatory Action of Glucocorticoids — New Mechanisms for Old Drugs, New England Journal of Medicine 353: 1711-1723.
14.Pockaj BA, Basu GD, Pathangey LB, Gray RJ, Hernandez JL, et al. (2004) Reduced T-cell and dendritic cell function is related to cyclooxygenase-2 overexpression and prostaglandin E2 secretion in patients with breast cancer. Ann. Surg Oncol 11: 328-339.
15.Misra S, Hascall VC, Markwald RR, O’Brien PE, Ghatak S (2018) “Inflammation and cancer” in Wound Healing: Stem Cells Repair and Restorations, Basic and Clinical Aspects, ed K. Turksen (John Wiley & Sons) 239-274.
16.Faden AI, Wu J, Stoica BA, Loane DJ (2016) Progressive inflammation-mediated neurodegeneration after traumatic brain or spinal cord injury. Br. J. Pharmacol 173: 681-691.
17.Gomez H, Ince C, De Backer D, Pickkers P, Payen D, et al. (2014) A unified theory of sepsis-induced acute kidney injury: inflammation, microcirculatory dysfunction, bioenergetics and the tubular cell adaptation to injury. Shock 41: 3.
18.Tucker PS, Scanlan AT, Dalbo VJ (2015) chronic kidney disease influences multiple systems: describing the relationship between oxidative stress, inflammation, kidney damage, and concomitant disease. Oxid. Med. Cell Longev 806358.
19.Langenbach R, Loftin C, Lee C, Tiano H (1999) Cyclooxygenase knockout mice: models for elucidating isoform-specific functions. Biochem. Pharmacol 58: 1237- 1246.
20.Kurumbail RG, Kiefer JR, Marnett LJ (2001) Cyclooxygenase enzymes: catalysis and inhibition. Curr. Opin. Struct Biol 11: 752-760.
21.Morita I (2002) Distinct functions of COX-1 and COX-2. Prostaglandins Other Lipid Mediat 68: 165-175.
22.Aid S, Langenbach R, Bosetti F (2008) Neuroinflammatory response to lipopolysaccharide is exacerbated in mice genetically deficient in cyclooxygenase-2. J. Neuroinflammation 5: 17.
23.Kankaanranta H, Moilanen E, Vapaatalo H (1991) Tolfenamic acid inhibits leukotriene B4-induced chemotaxis of polymorphonuclear leukocytes in vitro. Inflammation 15: 137-143.
24.Proudman KE, McMillan RM (1991) Are tolfenamic acid and tenidap dual inhibitors of 5-lipoxygenase and cyclo- oxygenase? Agents Actions 34: 121-124.
25.Moilanen E, Kankaanranta H (1994) Tolfenamic acid and leukotriene synthesis inhibition. Pharmacol. Toxicol 75: 60- 63.
26.Sidhu PK, Landoni MF, Lees P (2005) Influence of marbofloxacin on the pharmacokinetics and pharmacodynamics of tolfenamic acid in calves. J. Vet Pharmacol Ther 28: 109-119.
27.Sidhu P, Landoni M, Lees P (2006) Pharmacokinetic and pharmacodynamic interactions of tolfenamic acid and marbofloxacin in goats. Res Vet. Sci 80: 79-90.
28.Ray WA, Stein CM, Daugherty JR, Hall K, Arbogast PG, et al. (2002) COX-2 selective non-steroidal anti-inflammatory drugs and risk of serious coronary heart disease. Lancet 360: 1071-1073.
29.Wang SS, Mu RH, Li CF, Dong SQ, Geng D, et al. (2017) microRNA-124 targets glucocorticoid receptor and is involved in depression-like behaviors. Prog Neuropsychopharmacol Biol Psychiatry 79: 417-425.
30.Ahn KS, Noh EJ, Zhao HL, Jung SH, Kang SS, et al. (2005) Inhibition of inducible nitric oxide synthase and cyclooxygenase II by Platycodon grandiflorum saponins via suppression of nuclear factor-kappaB activation in RAW 264.7 cells. Life Sci 76: 2315-2328.
31.Srivastava KD, Kattan JD, Zou ZM, Li JH, Zhang L, et al. (2005) The Chinese herbal medicine formula FAHF-2 completely blocks anaphylactic reactions in a murine model of peanut allergy. J. Allergy Clin. Immunol 115: 171-178.
32.Shin CY, Lee WJ, Lee EB, Choi EY, Ko K H (2002) Platycodin D and D3 increase airway mucin release in vivo and in vitro in rats and hamsters. Planta Med 68: 221-225.
33.Azam F, Prasad M V, Thangavel N, Shrivastava A K, Mohan G (2012) Structure-based design, synthesis and molecular modelling studies of thiazolyl urea derivatives as novel anti- parkinsonian agents Med Chem 8: 1057-1068.
34.Shushni SMA, Azam F, Lindequist U (2013) Oxasetin from Lophiostoma sp. of the Baltic Sea: identification, in silico binding mode prediction and antibacterial evaluation against fish pathogenic bacteria Nat. Prod Commun 8: 1223-1226.
35.Harborne JB (1984) phytochemical methods - a guide to modern techniques of plant analysis. (Chapman and Hall, 2nd eds.) 4-16.
36.Karawya M S, Abdel-Wahab S M, Zaki A Y (1971) Colorimetric Method for the Estimation of Alkaloids in Lobelia and Its Pharmaceutical Preparations. Journal of Association of Official Analytical Chemists 54: 1423-1425.
37.Malick C P, Singh M B (1980) Estimation of polyphenols, In: CP. Malik, MB. Singh, (eds). In: Int. J. Adv Res. Bil.Sci 2: 185-189.
38.Makkar Robert E B (1971) Method for estimation of Tannins in grain sorghum. Agronomy Journal 63: 511.
39.Sadasivam S Manickam (2008) A Biochemical Method. 3rd Edition, New Age International Publishers New Delhi 9.
40.Madhu M, Sailaja V, Satyadev T N V S S, Satyanarayana M V (2016) Quantitative phytochemical analysis of selected medicinal plant species by using various organic solvents. Journel of pharmacognocy and phytochemistry 5: 25-29.
41.Senguttuvan J, Paulsamy S, & Karthika K (2014) phytochemical analysis and evaluation of leaf and root parts of the medicinal herb, Hypochaeris radicata L. for in vitro antioxidant activities. Asian Pacific Journal of Tropical Biomedicine 4: 359-367.
42.Paulsamy S, Jeeshna M V (2011) Preliminary phytochemistry and antimicrobial studies of an endangered medicinal herb Exacumbicolor Roxb. Res J Pharm Biol Chem Sci, 2: 447- 457.
43.Truong Dieu-Hien, Nguyen Dinh Hieu, Ta, Nhat Thuy Anh, Bui Anh Vo, Do Tuong Ha, et al. (2019) Evaluation of the Use of Different Solvents for Phytochemical Constituents, Antioxidants, and In Vitro Anti-Inflammatory Activities of Severinia buxifolia. Journal of Food Quality 29: 1-9.
44.Edoga H O, Okwu D E, Mbaebie B O (2005) Phytochemicals constituents of some Nigerian medicinal plants. Afr J. Biotechnol 4: 685-688.
45.Mann J (1978) Secondary Metabolism. London. Oxford University press 154.
46.Vasu K, Goud J V, Suryam A, Singara Chary M A (2009) Biomolecular and phytochemical analyses of three aquatic angiosperms. Afr. J. Microbiol Res 3: 418-421.
47.Muthu C, Ayyanar M, Raja N, Ignacimuthu S (2006) Medicinal plants used by traditional healers in Kancheepuram District of Tamil Nadu, India, Journal of Ethnobiology and Ethnomedicine 2: 43.
48.Inayatullah S, Prenzler P D, Obied H K, A-u Rehman, Mirza B (2012) Bioprospecting traditional Pakistani medicinal plants for potent antioxidants. Food Chem 132: 222-229.
49.Arunakumara KKU, Walpol BC, Subasinghe S, Min-Ho Yoon (2011) Pteroarpus Santalinuslinn.f. : A Review of its Botany, uses, Phytochemistry and Pharmacology. Journal of the Korean Society for Applied Biological Chemistry 54: 495-500.
50.Kondeti V K, Kameswara Rao B, Maddirala DR, Thur S K M, Fatima SS, Kasetti RB, et al. (2010) Effect of Pterocarpus santalinus bark, on blood glucose, serum lipids, plasma insulin and hepatic carbohydrate metabolic enzymes in streptozotocin-induced diabetic rats. Food Chem Toxicol 48: 1281-1287.
51.Narayan S, Devi R S, Devi CSS (2007) Role of Pterocarpus santalinus against mitochondrial dysfunction and membrane lipid changes induced by ulcerogens in rat gastric mucosa. Chem biol interact 170: 67-75.
52.Kesari A N, Gupta R K, Watal G, (2004) Two aurone glycosides from heartwood of Pterocarpus santalinus. Phytochemistry 65: 3125-3129.
53.Lacaille-Dubois M A, Wagner H (1996) A review of the biological and pharmacological activities of saponins. Phytomedicine 4: 363-386.
54.Francis G, Kerem Z, Makkar H P S, Becker K (2002) the biological action of saponins in animal systems: A review. Brit. J. Nutr 88: 587-605.
55.Petti S, Scully C (2009) Polyphenols, oral health and disease: a review. J. Dent 37: 413-423.
56.Petrovska B B (2012) Historical review of medicinal plants’ usage. Pharmacogn. Rev 6: 1-15.
57.Daniel M (2016) Medicinal Plants: Chemistry and Properties. CRC Press.
58.Schmelzer G H, Gurib-Fakim A (2008) Tabernaemontana elegans Stapf. In Schmelzer G H; (eds.) Medicinal Plants 1. Plant Resources of Tropical Africa. PROTA Foundation; Backhuys Publishers; CTA 11: 592-593.
59.Mairura F S, Schmelzer G H (2008) Tabernaemontana crassa Benth. In Schmelzer, GH Gurib-Fakim A (eds.) Medicinal Plants 1. Plant Resources of Tropical Africa. PROTA Foundation; Backhuys Publishers CTA 11: 589-592.
60.Ye?ilada E, Küpeli E (2002) Berberis crataegina DC. Root exhibits potent anti-inflammatory, analgesic and febrifuge effects in mice and rats. J. Ethnopharmacol 79: 237-248.
61.Danz H, Stoyanova S, Wippich P, Brattström A, Hamburger M (2001) Identification and isolation of the cyclooxygenase-2 inhibitory principle in Isatis tinctoria. Planta Med 67: 411- 416.
62.Anand B, Shrihari M (2010) Antioxidant and imrnuno modulatory activity of the alkaloidal fraction of Cissampelos pareira Linn. Sci Pharm 78: 21- 31.
63.Kuete V, Ango P Y, Fotso G W, Gilbert DWF Kapche, Berhanu M Abegaz, et al. (2011) Antimicrobial activities of the methanol extract and compounds from Artocarpus communis (Moraceae). BMC Complement Altern Med 11: 42.
64.Simão da Silva, Kathryn AB, Paszcuk Ana F, Passos Giselle F, Silva Eduardo S, et al. (2011) Activation of cannabinoid receptors by the pentacyclic triterpene α,β-amyrin inhibits inflammatory and neuropathic persistent pain in mice. Pain 152: 1872-1887.
65.Qing ZX, Huang JL, Yang XY, Jing-Hong Liu, Hua-Liang Cao, et al. (2018) Anticancer and reversing multidrug resistance activities of natural isoquinoline alkaloids and their structure-activity relationship. Curr Med Chem 25: 5088-5114.
66.Suh YG, Kim YH, Park M H, Choi Y H, Lee H K, et al. (2001) Pimarane cyclooxygenase 2 (COX-2) inhibitor and its structure-activity relationship. Bioorg. Med. Chem. Lett 11: 559-562.
67.Küpeli E, Ko?ar M, Ye?ilada E, Ba?er KHC (2002) a comparative study on the anti-inflammatory, antinociceptive and antipyretic effects of isoquinoline alkaloids from the roots of Turkish Berberis species. Life Sci 72: 645-657.
68.Kuo C L, Chi CW, Liu T Y (2004) the anti-inflammatory potential of berberine in vitro and in vivo. Cancer Lett 203: 127-137.
69.Fukuda K, Hibiya Y, Mutoh M, Koshiji M, Akao S, et al. (1999) Inhibition by berberine of cyclooxygenase-2 transcriptional activity in human colon cancer cells. J. Ethnopharmacol 66: 227-233.
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