Sains Malaysiana 49(1)(2020): 93-101

http://dx.doi.org/10.17576/jsm-2020-4901-11

Protective Effect of Cocoa Extract on Ethanol Induced Liver Injury in Sprague-dawley Rats

(Kesan Pelindung Ekstrak Koko untuk Kecederaan Hati Teraruh Etanol pada Tikus Sprague-dawley)

ROSMAWATI MAT SHAIR^1,2, MOHAMAD YUSOF MASKAT²*, MOHAMAD KHAN AYOB² & ROSMIN KASRAN¹

¹Division of Biotechnology, Cocoa Innovation and Technology Centre, Malaysia Cocoa Board, Nilai Industrial Park, 71800 Nilai, Negeri Sembilan Darul Khusus, Malaysia

²Center of Biotechnology & Functional Food, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor Darul Ehsan, Malaysia

Diserahkan: 19 Jun 2019/Diterima: 15 Oktober 2019

ABSTRACT

Cocoa is a rich source of dietary polyphenol, highly potential antioxidant against free radicals. This study was designed to identify the effect of cocoa polyphenol extract in protecting from ethanol-induced liver injury in rats. Fifty male Sprague-dawley rats were divided into five groups fed with or without ethanol (4 g/kg/d), cocoa extract (300 mg/kg/d) and silymarin (200 mg/kg/d) continuously for 3 weeks using an enteral feeding protocol. All treatments were given orally every day for three weeks and continuously supply food and water ad libitum. Results showed that cocoa extract (CE) from unfermented cocoa beans had a total polyphenol content of 335.70±27.51 mg GAE/g and 38.10±4.52 mg CaE/g. Meanwhile, analysis normal phase-high performance liquid chromatography shows CE contains 59.47±9.44 mg/g and 14.69±1.63 mg/g of epicathechin and catechin, respectively, which is three fold higher compared to commercial cocoa powder. It also contains 59.69±2.15 mg/g theobromine which also three fold higher compared to caffeine 19.87±1.37 mg/g. In vitro study showed cocoa extract contains high antioxidant activities by 91.9±1.00 % against superoxide scavenging system (O₂^-) and 97.7±0.15% against  a-a-diphenyl-β-picrylhydrazyl radical (DPPH) systems. In vivo study showed increasing level in both liver function enzymes, aspartase aminotransferase (AST) and alanine aminotransferase (ALT) in ethanol intoxication by 116.80±5.23 mmol/L and 56.37±2.71 mmol/L, respectively. Ethanol intoxication was blocked by cocoa extract nearly 89.95±1.18 mmol/L and 46.75±0.74 mmol/L, respectively, and it was comparable with SDT group for both enzymes AST and ALT by 112.19±6.02 mmol/L and 42.49±0.62 mmol/L, respectively. Furthermore, ethanol groups showed significantly lower (p<0.05) of glutathione level by 0.29 ±0.03 µmol/g, however cocoa extract with antioxidant defense system either direct or indirectly protect liver injury by increasing glutathione level at 0.53±0.02 µmol/g. As a result, cocoa extract shows its potential as antioxidant agents to protect ethanol-induced liver injury.  

Keywords: Antioxidant activity; chronic ethanol; cocoa extract; glutathione; liver injury

ABSTRAK

Koko kaya dengan sumber polifenol diet bertindak sebagai antioksida dan berpotensi untuk menyingkirkan radikal bebas. Kajian ini dilakukan bagi mengenal pasti kesan ekstrak polifenol daripada koko untuk mencegah kerosakan hati tikus yang disebabkan oleh pengambilan etanol. Sebanyak lima puluh ekor tikus Sprague-dawley jantan, dibahagikan kepada lima kumpulan iaitu kumpulan kawalan, etanol (4 g/kg/d), ekstrak koko (300 mg/kg/d) dan silimarin (200 mg/kg/d) yang mengandungi sepuluh ekor bagi setiap kumpulan. Setiap rawatan diberikan secara oral selama 3 minggu dan pengambilan makanan dan minuman adalah secaraad libitum. Analisis Folin-ciocalteau ekstrak koko (CE) telah menunjukkan bahawa jumlah kandungan polifenol ialah sebanyak 335.70±27.51 mg GAE/g dan 38.10±4.52 mg CaE/g. Manakala keputusan fasa normal kromatografi cecair berprestasi tinggi (NP-HPLC) pula menunjukkan ekstrak koko mengandungi kandungan epikatekin sebanyak59.47±9.44 mg/g dan katekin sebanyak 14.69±1.63 mg/g iaitu tiga kali ganda lebih tinggi berbanding serbuk koko komersial. Ia juga mengandungi theobromina sebanyak 59.69±2.15 mg/g, iaitu tiga kali ganda kandungan lebih tinggi berbanding kafein iaitu 19.87±1.37 mg/g. Kajianin vitro menunjukkan aktiviti antioksida ektrak koko adalah sangat tinggi iaitu sebanyak 91.9±1.00% terhadap radikal superoksida (O₂^-) dan 97.7±0.15% terhadap radikala-a-difenil-β-pikrilhidrazil (DPPH). Manakala kajianin vivo menunjukkan berlaku peningkatan terhadap kedua-dua enzim fungsi hati iaitu enzim aspartase aminotransferase (AST) dan alanine aminotransferase (ALT) di dalam ketoksikan etanol masing-masing sebanyak 116.80±5.23 mmol/L dan 56.37±2.71 mmol/L. Ketoksikan etanol dapat dicegah dengan kahadiran ekstrak koko dengan kandungan enzim masing-masing adalah sebanyak 89.95±1.18 mmol/L dan 46.75±0.74 mmol/L, selari dengan kumpulan silimarin bagi kedua-dua enzim AST dan ALT iaitu masing-masing 112.19±6.02 mmol/L dan 42.49±0.62 mmol/L. Selanjutnya, kumpulan etanol menunjukkan penurunan yang signifikan (P<0.05) kandungan glutation sejenis antioksida pertahanan badan semula jadi sebanyak 0.29 ±0.03 µmol/g, walau bagaimanapun, kehadiran ekstrak koko yang bertindak samada secara langsung atau tidak langsung melindungi kerosakan hati telah meningkatkan kandungan glutation sebanyak 0.53±0.02 µmol/g. Kesimpulannya, ekstrak koko telah menunjukkan bahawa ia sangat berpotensi sebagai agen antioksida dalam melindungi kerosakan hati yang disebabkan oleh pengambilan etanol.

Kata kunci: Aktiviti antioksida; ekstrak koko; etanol kronik; glutation; kerosakan hati

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