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Journal of Oleo Science Copyright ©2018 by Japan Oil Chemists’ Society J-STAGE Advance Publication date : November 15, 2018 doi : 10.5650/jos.ess18064 J. Oleo Sci. Role of Borage Seed Oil and Fish Oil with or without Turmeric and Alpha- Tocopherol in Prevention of Cardiovascular Disease and Fatty Liver in Rats Sahar Y. Al-Okbi1＊ , Salah M. El-qousy2, Samah El-Ghlban3, and Hosam F. Moawad4 1 Nutrition and Food Sciences Department, National Research Centre, Cairo, EGYPT 2 Chemistry Department, Faculty of Science El Menoufeia University, Shebin El Kom, EGYPT 3 Chemistry Department, Biochemistry Division, Faculty of Science El Menoufeia University, Shebin El Kom, EGYPT 4 Medical Research Center, Faculty of Medicine, Ain Shams University, Cairo, EGYPT Abstract: The aim of the present research was to Study the prevention of dyslipidemia, oxidative stress, inflammation and fatty liver as risk factors for cardiovascular disease via intervention by borage oil (B) and fish oil (F) with or without turmeric (T) and alpha-tocopherols (TC). Fatty acids were assessed in both oils while curcuminoids were determined in turmeric. Rats were divided into; first group fed on balanced diet and designated as normal control (NC), second fed on dyslipidemic and steatohepatitis (DS) inducer diet which represented the DS control group and groups 3-6 fed on DS inducer diet with daily oral administration of B, B+T+TC, F and F+T+TC; respectively for 5 weeks. Liver fat and plasma lipid profile, oxidative stress and inflammatory biomarker and liver and heart histopathology were assessed. Results showed gamma linolenic to be 21.01% in B. F contained eicosapentaenoic as 22.768% and docosahexaenoic acid as 13.574%.Total curcuminoids were 4.63 mg/g turmeric. The DS control group showed significant dyslipidemia, elevated malondialdehyde (MDA), tumor necrosis factor-alpha and liver fat with significant reduction in total antioxidant capacity (TAC) compared to NC. The different treatments produced significant improvement in all the parameters and histopathology. F was superior to B in ameliorating liver histopathological changes while B was more efficient in elevating TAC. B was more promising in improving lipid profile and liver fat compared to B + T + TC, while the latter was superior in improving MDA and liver histopathology. Fish oil was more efficient than F+TC+T except for TAC and high density lipoprotein cholesterol which were more improved on addition of TC and T. Conclusion: Borage and fish oil with or without antioxidants protect from cardiovascular and fatty liver diseases with variable degrees. Key words: borage oil, fish oil, turmeric, alpha-tocopherol, cardiovascular disease, fatty liver 1 Introduction omega3 and omega6 could have different efficiency Cardiovascular diseases（CVDs）are considered as the towards dyslipidemia and CVDs. Omega3 fatty acids like al- major cause of morbidity and death worldwide in both de- pha-linolenic, eicosapentaenoic（EPA）and docosahexaenoic veloped and developing countries1）. Dyslipidemia, oxidative （DHA）possess both hypolipidemic and anti-inflammatory stress and inflammation are among the reported risk activity but there is a confliction in literature concerning factors for cardiovascular diseases2）. Recently fatty liver their effect on oxidative stress4）. Fish oil is rich in long with inflammation（steatohepatitis）, which is increasing chain polyunsaturated fatty acids（EPA and DHA）and was worldwide, is accused as being major player in induction of reported to possess the aforementioned health effects as cardiovascular diseases3）. Generally, the condition includ- well as hepatoprotective effect5）. On the other hand little is ing dyslipidemia, inflammation, steatohepatitis, oxidative known about the effect of omega6 fatty acid like linoleic stress and impaired glucose tolerance is called metabolic acid on metabolic syndrome. Although linoleic was report- syndrome. ed to have hypolipidemic effect but claimed to induce some Two series of polyunsaturated fatty acids represented by sorts of pro-inflammation. However; this claim was not ＊Correspondence to: Sahar Y. Al-Okbi, Nutrition and Food Sciences Department, National Research Centre, Cairo, EGYPT E-mail: [email protected] Accepted August 20, 2018 (received for review April 8, 2018) Journal of Oleo Science ISSN 1345-8957 print / ISSN 1347-3352 online http://www.jstage.jst.go.jp/browse/jos/ http://mc.manusriptcentral.com/jjocs 1 S. Y. Al-Okbi, S. M. El-qousy, S. E.-Ghlban et al. supported by recent studies6, 7）. Linoleic acid was reported 2.2 Animals by Maruyama et al.8）to be a potent protector against lipo- Male Sprague Dawley rats weighing 140-150 g. were toxicity induced inflammation. Other omega6 fatty acid like used in the present study. Animals were obtained from gamma linolenic（GLA）was reported to have anti-inflamma- Animal house of National Research Centre, Cairo, Egypt. tory and antioxidant effect9, 10）. The ratio of n-6/n-3 polyun- Rats were kept individually in stainless steel cages; water saturated fatty acids is a better index of health benefit; un- and food were given adlibtium. Animal experiment was favourable ratio was reported to be associated by greater carried out according to the Medical Research Ethics Com- systemic inflammation11）. mittee for institutional and national guide for the care and Borago officinalis is native to the Mediterranean region, use of laboratory animals, National Research Centre, Cairo, North Africa and Middle East and is known as starflower or Egypt. borage. In traditional medicine borage is used in gastroin- testinal, respiratory, cardiovascular and urinary disor- 2.3 Methods ders12）. It is also used for pre and post-menopausal symp- 2.3.1 Assessment of fatty acids in borage and fish oil toms13）. Commercially, the plant is cultivated for borage Methyl esters were prepared via trans-esterification of seed oil which is used for different medical and nutritional fatty acids using methanol sulfuric acid and chloroform benefits. The seed contain 26-38％ oil which is the richest mixture according to Indarti et al.18）. Twenty mg of the oil source of gamma linolenic acid（GLA）that accounts for was weighed into clean, 10 mL screw-cap glass tubes, to 17-28％ of the oil14）. which 4 mL fresh solution of a mixture of freshly prepared Aside from being a popular condiment, turmeric has long methanol, concentrated sulfuric acid and chloroform history in medicinal use in Ayurvedic medicine for wound （1.7/0.3/2 v/v/v）was added. The tubes were covered; trans- healing, hepatic diseases, joint pains, gastrointestinal disor- esterification was run at 100℃ for 30 min. On completion der and cancer prevention. Curcuminoids and phenolic of the reaction, the tubes were brought to room tempera- compounds as well as the essential oil were reported to be ture. Then, 3 mL distilled water was added into the mixture the major bioactive constituents in turmeric and to which and thoroughly mixed by vortex for 1 min. After the forma- the bioactivity of turmeric were ascribed. The health bene- tion of two phases, the lower phase containing the fatty fits of such constituents are attributed mainly to their anti- methyl esters was transferred to clean glass tube and dried 15） oxidant and anti-inflammatory effect . A meta analysis with anhydrous Na2SO4. Fatty acids were analyzed by GC study showed curcuminoids therapy to reduce triglycerides after addition of a known volume of chloroform. and increase HDL-Ch16）. Alpha-tocopherol an antioxidant Assessment of the methyl ester was carried out by in- oil soluble vitamin was reported recently to possess anti- jecting 2 μL into a Hewlett Packard HP-system 6890 gas inflammatory, hepatoprotective and cardioprotective chromatograph equipped with FID. HP-5 capillary column effect17）. So it is hypothesized that combination of either （30 m×0.32 mm i.d.; 0.25 μm film thickness）was used to fish oil or borage seed oil with both turmeric and alpha to- separate the different methyl esters. The chromatographic copherol could impart an efficient cardiovascular protec- analysis conditions were; initial temperature 70℃ with a tive effect in addition to prevention of steatohepatitis. hold for 1 min, then raised to 120℃ at a rate of 40℃/min The objective of the present research was to set a com- with 2 min hold then the temperature was finally raised to parative study between the healthy effect of borage and 220℃ at a rate of 4℃/min with another 20 min hold. The fish oil in rat model of metabolic syndrome（dyslipidemia injector and detector temperatures were 250℃ and 280℃; and steatohepatitis）when administered alone or as a blend respectively. Identification of the fatty acid methyl esters with alpha tocopherol and turmeric. The aim included the were carried out by direct comparison of retention times of analysis of fatty acids contents of the oils and assessment each of the separated compounds with standards of the of curcuminoids content of turmeric. fatty acid methyl esters analyzed under the same condi- tions. Quantization was based on peak area integration. 2.3.2 Curcuminoids determination in turmeric Curcuminoids were determined according to Jayapraka- 2 Experimental sha et al.19）. Methanolic stock solutions of curcumin, deme- 2.1 Materials thoxycurcumin, and bisdemethoxycurcumin were prepared Borage oil（Borago officinalis, family Boraginaceae）and separately at a concentration of 0.5 mg/mL. Turmeric fish oil were obtained from Iherb, 1795 World Wide Blvd, powder sample（1.0 g）was extracted with hexane（50 mL） USA. Turmeric powder was purchased from herbal store, by using a Soxhlet extractor for 30min. The hexane extract local market, Cairo, Egypt. Alpha tocopherol was obtained was discarded, and the powder was re-extracted with 50 from local pharmacy, Cairo, Egypt; the manufacturer is mL of methanol for 2 h. One milliliter of this solution was Pharco Pharmaceuticals Company, Alexandria, Egypt. transferred to a 10 mL volumetric flask, and the volume was adjusted to 10 mL with methanol. 2 J. Oleo Sci.

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