Efficacy of Chia Seeds (Salvia hispanica L) on Serum Cardiovascular Risk Factors in Male Wistar Rats Fed a High Fat and Fructose Diet

Abstract

Cardiovascular disease (CVD) is the world’s biggest killer claiming about 17.9 million lives annually where 1/3 being prematurely under 70 years. The fundamental task of dealing with CVD epidemic is primary prevention of CVD risk factors using natural food sources to reduce side effects caused by synthetic drugs. Chia seeds (Salvia hispanica L) are among of the natural plant-based foods that contain the greatest amount of nutrients particularly omega-3 fatty acid (α-linolenic acid), fiber, protein, fats and minerals. They are considered a functional food with pronounced health benefits and deemed useful in cardiovascular health. The objectives of this study were to; determine the proximate composition, mineral contents, fatty acid profiles and phytochemical screening of chia seeds grown in East Africa (Kenya and Uganda); investigate the in vitro efficacy of chia seed extracts on the inhibition of α-amylase and pancreatic lipase activities; determine the effect of ground chia seeds/extracts on postprandial glycaemia, body weight, hematological parameters and cellular morphology in rats; and evaluate the dose effect of chia seed extracts on plasma glycaemia, body weight changes, lipemia, liver enzymes and hematological parameters in male Wistar rats fed with fructose and lard. Samples, black chia Among’ura-Teso (BCAT), black chia Kigumba (BCK), black chia Molo (BCM), Black chia Ongariama-Teso (BCOT) and White chia Bukembo (WCB) were collected from Kenya and Uganda. Official methods of analysis (AOAC International) were adopted for proximate analysis; minerals were determined by Atomic absorption spectrophotometer, and fatty acid profiles were analyzed by Gas chromatography. Phytochemicals and enzymatic assays for pancreatic lipase and α-amylase were conducted using standard methods. Acarbose and Orlistat were used as positive controls for enzymatic assays. For animal studies, 44 male Wistar rats were divided into two experimental groups (acute experiment-20 rats and dose effect experiment-24 rats). In acute experiment, 20 male Wistar rats were assigned into three experimental groups and a control (n =5). Each experimental group received 10 g/20 g fructose/lard. Group 1 and 3 were supplemented with 20 g chia seed extract while group 2 received ground chia seeds only. Control group received rat pellets only. Each rat received 15g of pellets while water were given ad libitum in all groups. All diets were fed for a period of 28 days. In a dose effect experiment, 24 male Wistar rats were randomly assigned into three experimental groups (low dose, medium dose and high dose of 6 animals each) and a control. The control group received 6g/12g fructose/lard; the experimental groups were given 6g/12g of fructose/lard and in addition received 12g/kg, 18 g/kg and 24 g/kg body weight of chia seed extract for low dose, medium dose and high dose respectively. Rat pellets (15g) were given to each rat while water was given ad libitum to all groups. The diets were fed daily for a period of 8 weeks. Statistical analysis was conducted using STATA version 14.1, a one-way analysis of variance (ANOVA) used Bonferroni multiple comparison test to determine the variability between groups, and significance was accepted at p ≤ 0.05. Results on proximate composition indicated high contents of protein, fat and fiber. The fatty acid profiles revealed great amounts of α-linolenic acid (ALA) (45.29-56.99 %) followed by linoleic acid (15.9-20.28%) and oleic acid (6.88-11.58%). Although the amount of ALA was high compared to other fatty acids analyzed in all samples, the difference was not significant (p = 0.7391). Mineral determination (mg/100g) displayed high contents of potassium (492.96-862.98), phosphorous (486.45-569.45), calcium (297.47-429.09) and magnesium (192.22-202.97) while considerable amount was observed for iron, zinc, manganese, and copper. There was a significant difference (p = 0.0001) in mineral content between BCK and WCB with exception of phosphorus. For the in vitro studies, BCAT displayed the highest inhibition on α-amylase activity with the IC50 value of 104.51 ± 0.24 μg/ml and 72.06 ± 0.12 μg/ml for ethanolic and methanolic extracts respectively, however they were significantly lower (p = 0.0001) than that of acarbose. On the other hand, BCAT exhibited the strong pancreatic lipase inhibitor with IC50 value of 90.02 ± 0.17 μg/ml and 96.19 ± 0.20 μg/ml) for ethanolic and methanolic extracts respectively, followed by BCM and BCK. Orlistat indicated a significantly higher (p = 0.001) IC50 (54.14 ± 0.15 μg/ml), establishing its relative potency as pancreatic lipase inhibitor than chia seed extracts. In an acute experiment, there was an increase in postprandial blood glucose levels in group 1 from week I to IV contrary to groups supplemented with chia seeds/extract indicating the ability of chia seeds in controlling blood glucose. Complete blood counts showed a significant increase (p = 0.008) in mean corpuscular hemoglobin concentration, basophils (p = 0.035), and platelets (p = 0.0001) in experimental groups compared to control. In a dose effect experiment, platelets were significantly lower (p = 0.001) in low dose group compared to a control and high dose groups. For liver enzymes, Gamma glutamyl transferase (GGT) decreased significantly (p = 0.02) in low and medium dose groups and bilirubin was significantly lower (p = 0.005) in experimental groups compared to a control. Serum lipids such as high density lipoprotein cholesterol (HDL-C) increased significantly (p = 0.041) in high dose group than the control while low density lipoprotein cholesterol (LDL-C) decreased significantly (p = 0.035) in high dose group as compared to control and medium dose groups (p = 0.021). The East African grown chia seeds, exhibited high amount of nutrients and fatty acids. The seeds inhibited the pancreatic lipase and α-amylase activities, these enzymes are responsible for the digestion of fats and carbohydrates respectively. Inhibition of these enzymes may help decrease hyperglycemia and hyperlipidemia the risk factors for CVD. The positive impact showed on LDL-C, HDL-C and triglycerides in rats fed on diet high in fat and fructose suggest the potential of chia seeds in reducing serum CVD risks. Generally, these results highlight that chia seeds could be among the preferred natural food sources for the primary prevention of CVD. Therefore, we suggest its moderate incorporation in diets as a healthy food ingredient.