Anti-inflammatory activities of aqueous and ethanolic extracts from anoectochilus burmannicus

ฤทธ์ิต้านการอักเสบของสารสกัดน้ำและเอทานอลจากนกคุ้มไฟ (Anoectochilus burmannicus)

Name: Phatcharaporn Budluang

ThaSH: Orchids -- Micropropagation

Classification :.DDC: 584.4

ThaSH: Anti-inflammatory agents

Abstract: Inflammation is a complex biological response of human body tissues to generally protect our body from injury and infection; however, chronic inflammation which is mediated by free radicals could lead to serious adverse effects including cancer, metabolic syndromes, and diabetes mellitus. Currently, a number of studies have demonstrated the obesity-induced chronic local inflammation in adipose tissue, which is also triggered by macrophages. Subsequently, several inflammatory cytokines, especially tumor necrosis factor-α (TNF-α) could induce the insulin resistance in adipocytes. Prevention or treatment of insulin resistance would be beneficial to diabetic patients. A variety of medicinal plants with anti-diabetic activity have been used as alternative medicine. Anoectochilus species such as Anoectochilus roxburghii and Anoectochilus formosanus have exerted both anti-inflammation and anti-insulin resistance effects. Yet, the bioactivities of Anoectochilus burmannicus (AB) remain unknown. This study therefore aimed to determine whether AB inhibits inflammation induced by lipopolysaccharide (LPS) and TNF-α in Raw 264.7 macrophages and 3T3- L1 adipocytes, respectively. Whole plant extract of AB was prepared using hot water or 80% ethanol. The yields of crude AB aqueous extract (ABAE) and AB ethanolic extract (ABEE) were 16.79% and 17.37%, respectively. Phytochemical composition analysis revealed more total phenolic contents in ABAE (19.09±1.18 mg/g extract) than ABEE (6.82±0.62mg/gextract). Using HPLC, several phenolic derivatives were found that ABAE has more contents of vanillic acid, coumaric acid and ferulic acid, but less content of chlorogenic acid than ABEE. ABAE and ABEE exhibited free radical scavenging effect with SC50 determined by DPPH assay at 4,551 ± 357 and 7,506 ± 832 μg/mL, respectively, and SC50 by ABTS assay at 421 ± 30 and 834 ± 58 μg/mL, respectively. Next, cytotoxicity of the extracts was determined using WST-1 assay, their anti-inflammatory activity using LPS-induced RAW 264.7 macrophages, and the anti-insulin resistant activity using TNF-α-treated 3T3-L1 adipocytes. The results showed that the extracts at the concentration up to 200 μg/mL were not toxic to both RAW 264.7 and 3T3-L1 cells. ABAE and ABEE significantly inhibited the nitric oxide (NO) production in LPSactivated macrophages in a dose-dependent manner. Moreover, ABAE (200 μg/mL) markedly reduced the expression of inducible nitric oxide synthase (iNOS) protein, and slightly decreased its mRNA expression in LPS-stimulated Raw 264.7 cells. Unlike ABAE, ABEE at lower dose (50 μg/mL) significantly inhibited the LPS-stimulated expression of iNOS at both protein and mRNA levels. In addition, ABAE slightly, while ABEE significantly reduced the LPS-induced protein expression of proinflammatory cytokine, interleukin-6 (IL-6). Both extracts significantly inhibited the LPS-induced mRNA expression of IL-6 and interleukin-1β (IL-1β) but not TNF-α. ABEE significantly decreased both mRNA and protein expression of cyclooxygenase-2 (COX-2), whereas ABAE slightly decreased the mRNA, but not protein expression in LPS-stimulated RAW 264.7 macrophages. Next, the model of inflammation-induced insulin resistance in obesity was performed in vitro by treating 3T3-L1 adipocytes with TNF-α leading to the reduction of glucose uptake by approximately 50%. ABAE and ABEE (50 and 200 μg/ml) significantly improved the insulin response of 3T3L-1 by increasing the cellular uptake of glucose. Furthermore, these extracts showed no cytotoxicity to normal human peripheral mononuclear cells (PBMCs) suggesting the extracts might be safe for further investigation in animal and clinical studies. In conclusion, the present study demonstrated apparently for the first time biological effects of the ABAE and ABEE against oxidative stress, inflammation and inflammation-induced insulin resistance. Inhibition of inflammation together with reduction of inflammation-stimulated insulin resistance by A. burmannicus extracts would be scientifically support usage and development of the extracts as folk medicine, food ingredient or food supplement for prevention of inflammation-related chronic diseases including diabetes mellitus type 2. Further studies should investigate the bioactive components in ABE and to provide an insight into the accuracy mechanisms of these actions.

Chiang Mai University Library

Address: Chiang Mai

Email: cmulibref@cmu.ac.th

Name: Piya Temviriyanukul

Role: Chairman

Name: Teera Chewonarin

Role: Member

Name: Pornsiri Pitchakarn

Role: Member

Name: Arisa Bonness

Role: Member

Created: 2016

Modified: 2017-10-24

Issued: 2017-10-24

วิทยานิพนธ์/Thesis

application/pdf

CallNumber: 584.4 P536A

CallNumber: 584.4 P536A

eng

DegreeName: Master of Science

Level: Master's Degree

Descipline: Biochemistry

Grantor: Chiang Mai University

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