Antilipidemic activity of organic solvent extract from Sorghum bicolor on rats with diet-induced obesity

Abstract

The present study evaluated the antiobesity and lipid-lowering effects from dichloromethane and ethyl acetate extracts of Hwanggeumchal Sorghum varieties on Sorghum bicolor. The Hwanggeumchal Sorghum ethyl acetate extracts significantly reduced the plasma total cholesterol and triglyceride levels significantly when given orally at a dose of 50 and 300 mg/kg/day to the high-fat diet-induced obese rats for 2 weeks. These findings demonstrate the excellent pharmacological potential of Hwanggeumchal Sorghum varieties to prevent obesity.

Introduction

A large percentage of people worldwide (in excess of 50% in the US and Russia, for example) are overweight, with a body mass index (BMI) between 25 and 29.9 kg/ m⁻². ^1,2 There is an increasing demand by patients to use natural products with antiobesity activity, due to the side effects associated with the use of insulin and oral hypoglycemic agents. On the other hand, plant products are generally considered to be less toxic with fewer side effects than synthetic products. Consequently, plant-derived materials have received increased attention as biochemical active agents in antihyperglycemia and antihyperlipidemia therapies. The study of such medicines might offer a natural key to unlock the obesity researcher (include to diabetologist) pharmacy for the future. So, some herbal (and/or supplements) drugs are a good source of natural antiobesity (including antidiabetic) agents. ³ The regulation of energy homeostasis for metabolic diseases is one of the most rapidly advancing subjects in biomedical research today. Breakthroughs in understanding of the molecular mechanisms regulating body weight have also provided potential opportunities for therapeutic intervention and brought renewed hope and vitality for the development of antiobesity drugs. ⁴ Sorghum bicolor L. Monech (Gramineae) is a drought-resistant low-input cereal crop grown throughout the world and can be an alternative source of oil having clinical advantages. Genus sorghum includes many species and subspecies, including grain sorghum, grass sorghum, sweet sorghum and broomcorn. It is used as food, animal feed, fibers as in wall board, fences, biodegradable packing material and for ethanol production. ⁵ Sorghum is an important food for people living in the semi-arid tropical areas of Africa and Asia. ⁶ Sorghum flour is rich in phytochemicals with a potential to impact human health in a beneficial manner. ⁷ The storage proteins of sorghum constitute 50%−60% of the total protein of the grain and have been classified into three main groups, according to their molecular weight, extractability and structure. Phenolic compounds in sorghum occur as phenolic acids, flavonoids and condensed. ^8,9 Condensed tannins (proanthocyanidins) occur in sorghums with a pigmented testa, which have dominant B1B2 genes. The tannins in sorghums have the highest levels of antioxidants of any cereal analyzed. ¹⁰ Sorghum tannins are 15−30 times more effective at quenching peroxyl radicals than simple phenolics; thus, they are potential biological antioxidants. Despite their possible beneficial effects as antioxidants, tannins have been linked to reduced protein digestibility of sorghum, because they bind with proteins and inhibit enzymes. ¹¹ The present study evaluated the lipid-lowering effects from Hwanggeumchal Sorghum dichloromethane (HSDM) and ethyl acetate (HSEA) extracts of Hwanggeumchal Sorghum (HGS) commonly cultivated in Korea as well as to give animal experiments about their antiobesity effect. To the best of our knowledge, this is the first report on the antiobesity effect of HGS extracts.

Material and methods

Plant materials and sample extracts for animal experiments

Sorghum cultivars were provided by the Department of Functional Crop, National Institute of Crop Science, Rural Development Administration, South Korea. Voucher herbarium specimens were deposited with the reference number (KNICS-579) in the Herbarium of the Department of Functional Crop. The botanical identification was made by one of the authors, Dr Ill-Min Chung of Kon Kuk University (Seoul, South Korea). The seeds were stored at 4°C. Sorghum cultivars used in this study was HGS. The HGS was ground and shade dried, coarsely powdered plant material (200 g) was defatted with n-hexane in a Soxhlet apparatus. The defatted plant material was successively refluxed three times (12 h, 24 h, 48 h) with 99.5% dichloromethane and 98% ethyl acetate extracts solution (20-fold) and then filtered through a Whatman No. 42. The HSDM and HSEA extracts were gathered and the ethanol was evaporated under reduced pressure at 45°C in a rotary vacuum evaporator (Buchi R II, Buchi, Switzerland), followed by lyophilisation. The stock solutions were kept at 4°C in the dark until further experiments.

Animals

Male Wistar rats (weight, 160 ± 15 g) were purchased from Orient Bio Inc (Sungnam, South Korea) and maintained under standard environmental conditions, with free access to water. All experiments conducted in this study were approved by the Animal Care Committee of the Wonkwang University School of Medicine. They were grouped as described by Hajhashemi and Abbasi. ¹² Three groups (control, HSDM and HSEA treated; n = 8) were used to study the effect of HSDM and HSEA on the serum lipid concentrations of normolipidemic rats (Table 1). Both groups were maintained on a standard diet for 2 weeks. Rats in the HSDM and HSEA-treated group were orally administered the extracts (50 and 300 mg·kg^–1·d^–1 for 2 weeks). We also assessed the effect of HSDM and HSEA on the serum lipid concentrations in 5 groups of rats (n = 8) maintained on a high-cholesterol diet (normal diet supplemented with 2% cholesterol and 0.5% cholic acid) for 2 weeks (Table 2). Group I was the control group. Groups II and III orally received HSDM and HSEA at doses of 50 and 300 mg·kg^–1·d^–1, respectively. Group V orally received clofibrate (100 mg·kg^–1·d^–1). The vehicle (isotonic saline), HSDM, HSEA or clofibrate was administered for 2 weeks. After this period, the animals were killed, and serum samples were obtained and maintained frozen at −20°C until further analysis.

Table 1.

Effect of organic solvent extracts from Hwanggeumchal Sorghum on the serum lipid concentrations of rats maintained on a normal diet ^a

Groups	TC (mg/dL)	LDL-C (mg/dL)	HDL-C (mg/dL)	TG (mg/dL)
Control	79.2 ± 1.7	21.3 ± 1.5	42.6 ± 7.2	73.5 ± 6.8
HSDM treated (300 mg·kg–1·d –1)	78.3 ± 1.3	18.4 ± 3.2	43.5 ± 6.7	72.5 ± 7.1
HSEA treated (300 mg·kg–1·d –1)	79.1 ± 1.3	17.4 ± 2.6	44.5 ± 6.1	74.3 ± 2.4

^a The values are the mean ± SD.

Table 2.

Effect of organic solvent extracts from Hwanggeumchal Sorghum on the serum lipid profiles of hyperlipidemic rats ^a

Groups	Dose (mg·kg–1·d–1)	TC (mg/dL)	LDL-C (mg/dL)	HDL-C (mg/dL)	TG (mg/dL)
Control	0	109.3 ± 3.4	49.3 ± 3.6	38.3 ± 5.3	111.2 ± 4.5
HSDM treated	50	108.1 ± 3.2	47.2 ± 5.7	43.1 ± 6.2	105.2 ± 5.9
	300	101.3 ± 3.7	45.6 ± 4.6	49.4 ± 1.5	102.4 ± 3.6
HSEA treated	50	95.4 ± 2.4 b	31.4 ± 2.5 b	45.7 ± 3.6 b	82.4 ± 2.5 c
	300	75.3 ± 2.9 c	11.3 ± 4.7 c	59.0 ± 4.2 b	61.4 ± 6.3 c
Clofibrate treated	100	98.2 ± 7.9 b	45.2 ± 4.1	42.3 ± 3.7	86.3 ± 5.6 b

Abbreviations: HSDM: Hwanggeumchal Sorghum dichloromethane, HSEA: Hwanggeumchal Sorghum ethyl acetate.

^a The values are the mean ± SD.

^b p < 0.01compared to the control group.

^c p < 0.001 compared to the control group.

Toxicity studies

Toxicity studies were carried out on normal healthy rats. ¹³ The HSDM and HSEA (300 mg/kg/day) were administered orally for 4 days of two groups of rats (n = 6) and the animals were kept under observation for mortality as well as any behavioral changes.

Determination of serum lipid levels

The serum total cholesterol (TC), triglyceride (TG) and high-density lipoprotein-cholesterol (HDL-C) levels were colorimetrically measured using commercially available kits (Asan Pharmaceutical, South Korea). ^4,14,15 The low-density lipoprotein-cholesterol (LDL-C) levels were estimated using Friedewald’s (1972) formula: LDL-C = TC − [HDL-C + (TG/5)]. The atherogenic index was calculated as (TC − HDL-C)/HDL-C. ¹⁶

Statistical analysis

The data were statistically analyzed by a one-way analysis of variance, followed by Duncan’s test. The results are presented as the mean ± standard deviation (SD).

Results and discussion

The HSDM and HSEA yielded 4.3% and 3.8% (v/w), with a pleasant smell. The HSDM and HSEA (300 mg/kg/day) have no signification toxicity from rats. HSEA dose-dependently reduced the serum TC, LDL-C and TG levels of the test rats. The Lipid Research Clinics Primary Prevention Trial has indicated that the plasma LDL-C concentrations and the risk of coronary artery disease are positively correlated. ¹⁷ The fact that HSEA significantly lowered the serum TC, LDL-C, and TG levels of our test animals indicates that it is a promising protective agent against coronary artery disease. Drugs such as statins lower the serum TC and LDL-C levels by inhibiting the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, a rate-limiting enzyme in cholesterol biosynthesis. ¹⁸ However, the mechanism underlying HSEA antiobesity effects is unclear, and further experiments are underway to elucidate this. Several studies reveal that elevated serum HDL-C levels are associated with a decreased risk of coronary disease. ¹⁹ Besides lowering the TC, LDL-C and TG levels, HSEA significantly increased the HDL-C levels and may therefore be an effective agent for the prevention of obesity and the management of coronary artery disease. ²⁰ Additional experiments are required to clarify the mechanism underlying HSEA antiobesity activity.