Chemical and biological studies of medicinal plants used by Chungtia villagers of Nagaland and Aboriginal people of New South Wales

This PhD study was based on the ethnomedicinal knowledge of the Chungtia Indigenous community of Nagaland, India and Aboriginal people of New South Wales (NSW), Australia. It follows former investigations of the Macquarie University’s Indigenous Bioresources Research Group on first hand documentation of medicinal plants used by the Chungtia villagers and first hand and published accounts of medicinal plants used by Aboriginal people of NSW for the treatment of skin related ailments including sores,wounds and skin infections. The overall objective of this study was to conduct chemical and biological investigations on medicinal plants used for the treatment of skin related conditions by the Chungtia villagers of Nagaland and Aboriginal people of NSW. A comprehensive literature review on 135 Chungtia medicinal plants documented by firsthand interviews was conducted. In addition, an updated literature review on plants used for the treatment of sores, wounds and skin infections was performed. The first hand information of the medicinal plants was compared with published reports of ethnobotanical and ethnomedicinal uses as well as chemical and biological studies worldwide. The review identified eleven medicinal plants used for the treatment of skin related diseases with none or limited reports of chemical and/or biological studies. These therefore have potential for further studies. One of these plants, Erythrina stricta Roxb. (Fabaceae), used for the treatment of skin infections, eczema and contact dermatitis by Chungtia villagers, was selected for detailed chemical and biological investigations. Bark of E. stricta was sequentially extracted with n-hexane, dichloromethane, ethyl acetate, methanol and water and the extracts were assayed against the bacterial strains methicillin sensitive Staphylococcus aureus, methicillin resistant S. aureus, multidrug resistant S. aureus, antibiotic sensitive Escherichia coli and Pseudomonas aeruginosa and the fungal strain Candida albicans. The most significant antimicrobial activity was observed with the dichloromethane, n-hexane and ethyl acetate extracts, with MIC values of 7.81μg/mL, 125 μg/mL and 125 μg/mL against a methicillin sensitive strain of S. aureus. The extracts were also screened for antioxidant activity by DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical scavenging and ferric reducing antioxidant power (FRAP) assay methods. All the extracts showed positive responses to the antioxidant assays. GC-MS analysis of the n-hexane extract identified twelve compounds, including the bioactive compounds caryophyllene oxide (16.31%), β-caryophyllene (9.06%), β-selinene (7.06%),α-selinene (6.86%), selin-11-en-4-α-ol (6.80%), α-copaene (3.31%) and α-eudesmol(4.60%). These compounds have been reported for having antimicrobial (β-caryophyllene,caryophyllene oxide, α-eudesmol, α-selinene, β-selinene, α-copaene and δ-cadenine) and anti-inflammatory (α-copaene, β-caryophyllene, caryophyllene oxide and δ-cadenine) activities. Fractionation of the dichloromethane extract by normal phase silica gel column chromatography yielded eight active fractions that showed antibacterial activity against the methicillin sensitive strain of S. aureus with MIC values ranging from 15.6 μg/mL to 1mg/mL. Purification of the active fractions using different chromatographic techniques ledto the isolation of seven antimicrobial and antioxidant compounds; erynone, wighteone,alpinum isoflavone, luteone, obovatin, erythrinassinate B and isovanillin. Erynone wasidentified as a novel compound and this is the first report of isolation of luteone, obovatinand isovanillin from the genus Erythrina and the first report of isolation of wighteone and erythrinassinate B from E. stricta. A literature review of medicinal plants used by Aboriginal people of NSW for skin related ailments was conducted and thirty two plants were identified with none or limited reports on chemical and biological studies. Following this review, Acacia falcata, Acacia implexa, Cassytha glabella, Eucalyptus haemastoma, Hibbertia scandens, Smilax glyciphylla,Sterculia quadrifida and Syncarpia glomulifera were selected for chemical and biological studies. 70% aqueous ethanolic extracts obtained from the eight selected species were screened for antibacterial and antioxidant activities. Antibacterial activity assays of the extracts were conducted against methicillin sensitive, methicillin resistant and multidrug resistant strains of S. aureus, E. coli and P. aeruginosa using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] microdilution assay. All the extracts apart from S. glyciphylla and S. quadrifida possessed antibacterial activity against all three strains of S. aureus. S. glyciphylla showed activity against only methicillin sensitive S. aureus and S. quadrifida did not show any activity against any bacterial strains. S. glomulifera was identified with having the most active plant extract with an MIC of 7.81μg/mL, followed by E. haemastoma with MIC 62.5 μg/mL and A. implexa with MIC 125μg/mL. Qualitative and quantitative phytochemical screening of the extracts was also conducted. This is the first report of antibacterial activity of A. falcata, A. implexa, C.glabella, E. haemastoma, H. scandens and S. glomulifera against methicillin resistant strains of S. aureus. Antioxidant activity of the eight NSW plant extracts was evaluated by DPPH free radical scavenging, ABTS [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)] radical scavenging activity and FRAP assay methods. E. haemastoma, A. falcata and A. implexa possessed high antioxidant activity, with IC50 values of 51.99 ± 1.17 μg/mL, 130.20 ± 5.37and 217.03 ± 3.80, respectively. Qualitative phytochemical screening identified the presence of terpenoids, flavonoids, steroids, alkaloids, saponins, tannins and anthraquinone glycoside classes of compounds within the extracts. The highest amount of total phenolic and condensed tannin contents were found for E. haemastoma, followed by A. implexa. Acacia falcata contained the highest amount of total flavonoid content, with E. haemastoma and C. glabella also containing a high flavonoid content. A significant correlation was observed between the antioxidant properties and the total phenolic and flavonoid contents, which suggested that the phenolic and flavonoid type of compounds present in the extracts are the major contributors to their antioxidant properties. This is the first report of antioxidant activity studies for A. falcata, A. implexa, C. glabella, E.haemastoma, H. scandens, S. quadrifida and S. glomulifera. The 70% aqueous ethanolic extract of S. glomulifera was partitioned with n-hexane, dichloromethane, ethyl acetate, n-butanol and water. The partitions were screened for antibacterial activity against methicillin sensitive, methicillin resistant and multidrug resistant strains of S. aureus, E. coli and P. aeruginosa. The n-hexane partition showed the greatest activity with an MIC of 7.81 μg/mL against all three strains of S. aureus. GCMS analysis of the n-hexane partition identified twenty-four phytochemicals that included the well-known antimicrobial, antioxidant and anti-inflammatory compoundsα-phellandrene, aromadendrene, α-copaene, geranial, globulol, terpinene-4-ol and spathulenol. The identification of antibacterial and antioxidant activities and bioactive constituents from E. stricta and the NSW medicinal plants provides support for their traditional medicina luses by the Chungtia community of Nagaland and Aboriginal communities of NSW for the treatment of skin related ailments and increases the knowledge on these relatively unexplored plants.