THE INFLUENCE OF CINNAMON AND GINGER ON SERUM GLUCOSE, TRIGLYCERIDES AND CHOLESTEROL FRACTIONS IN ALBINO INDUCED DIABETIC RATS

Introduction

Diabetes mellitus (DM) is a heterogeneous metabolic disorder, a high occurrence of disease in noted and the numbers of diabetic patients are gradually increasing, thus the disease constitutes a major health concern(Narendhirakannan et al., 2005 and Etuk, 2010).

According to the World Health Organization (WHO), there are approximately 347 million diabetics worldwide, the number of diabetics had been double in the last few years and diabetes death will increase by two thirds between 2008 and 2030 (WHO, 2012). Reasons for this rise include an increase in sedentary lifestyle, the consumption of energy-rich diet, obesity (Yajmk, 2001). Diabetes is the fourth or fifth leading cause of death in most high-income countries and there are substantial evidences that it is epidemic in many economically developing and newly industrialized countries (IDF, 2011).

Plants have always been an exemplary source of drugs, it’s used in traditional medicine to treat diabetes mellitus represent a valuable alternative for the management of this disease, amongst such plants reported to have beneficial effects in the treatment of diabetes are spices such as cinnamon and ginger (Ugwuja et al., 2010; Gandhi and Sasikumar, 2012).

Cinnamon is a common spice used by different cultures around the world for several centuries. It is obtained from the inner bark of trees from the genus Cinnamomum, a tropical evergreen plant that has two main varieties; Cinnamomum zeylanicum (CZ) and Cinnamon cassia (CC) (also known as Cinnamomum aromaticum / Chinese cinnamon). In addition to its culinary uses, in native Ayurvedic medicine Cinnamon is considered a remedy for respiratory, digestive and gynecological ailments. Almost every part of the cinnamon tree including the bark, leaves, flowers, fruits and roots, has some medicinal or culinary use. The volatile oils obtained from the bark, leaf, and root barks vary significantly in chemical composition, which suggests that they might vary in their pharmacological effects. (Shen et al.,  2002).

Ginger (Zingiber officinale, Roscoe Zingiberaceae) is one of the most widely consumed spices for the flavoring of food worldwide (Li et al., 2012). Ginger rhizome has been used in traditional herbal medicine. Ginger has enormous health promoting potential effects in number of ailments including degenerative disorders (arthritis and rheumatism), digestive health (indigestion and constipation), cardiovascular disorders (atherosclerosis and hypertension), diabetes mellitus and cancer. Also it has anti-inflammatory properties, which are beneficial in controlling the process of aging. Moreover, it has antimicrobial potential, which can help in treating infectious diseases and helminthiasis (Jiang et al., 2006; White, 2007; Ali et al., 2008 and Nicoll and Henein, 2009).

This investigation was carried out in an attempt to study the influence of cinnamon and ginger on serum glucose, triglycerides and cholesterol fractions in diabetic rats.

Materials and Methods

Materials

Source of samples

5kg of both cinnamon and ginger were obtained from Local Market, Assiut, Egypt, in March (2014).

Preparation of samples

Preparation of (5% and 10%) concentration of cinnamon and ginger (50 and 100 g) were dissolved in 10 ml toleween 80 (obtained from El Gomhorya Company) then diluted to 500 ml with distilled water, then were shacked for 6 hours.

Experimental animals

One hundred and ten adult male white albino rats (Sprague dawley strain) weighing between (120 g±5) were obtained from the animal house of the Faculty of Medicine, Assuit University. The animals were housed as groups in wire cages under the normal laboratory individually conditions and fed on the basal diet the rats were fed for a week as adaption period. Body weight gain and feed intake weighed weekly and at the end of the experimental feeding period.

Basal diet and untreated diabetic diet

The basal diet used is outlined in table (1)

Table (1): Constituents of the basal diet for 100g diet.

Reeves et al., (1993).

Chemicals

Alloxan (Sigma, chemical company Lot 110H3367 for laboratory use only) was pruched from New Assiut Company, Toleween 80 was obtained from El Gomhorya Company and Atorvastatin + Gliclagide were obtained from local pharmacy, Assiut city, Assiut.

Methods

Design of the experiment

The rats were randomly allocated into (9) main Groups of (10) rats each. The number of animals in need during the course of the study was collectively (90) males the other rest (20) were used to overcome the loss in number of animals due to sudden death. Each rats was marked on the tail to differentiate between the animals in the (9) groups. Daily administrations were continued for two successive, periods (6) weeks each. In the first period group used as control and was fed on basal diets while the other eight groups were injected intraperitoneal with alloxan in a single does of 150 mg/kg body weight (Pang et al,. 1985). The drug was dissolved in distilled water. In the eight groups the animals were tested for diabetes after five days the start of injection of alloxan . The animals were considered diabetic when its glucose level was 250 mg/100 ml or more (normal blood glucose level was ranged between 90-120 mg/100 ml)

After the onset of induced diabetic with alloxan, the animals were reclassified into the following groups fed on treated cinnamon and ginger as follows:

Treated cinnamon and ginger groups

Ginger and cinnamon extract were given orally to the rats through a gastric tube daily at a dose of 4 ml/kg body weight.

Blood sampling

At the end of each experiment, rats were fasted overnight and anesthetized. Blood samples were collected from the retro-orbital plexus from all animals of each group into clean, dry and labeled tube. The tubes contained heparin (10.0 IU/ml) as anticoagulant. Blood was centrifuged (3500 r-p. m for 15 min) to separate plasma. Which was tightly kept in sealed aliquot tubes at 20°C until biochemical assays.

Biochemical methods

          Prepared samples (as mentioned in blood sampling) were used to study the following biochemical parameters using PHOTO Mech 301-D spectrophotometer (Optima).

Determination of blood serum glucose

Blood serum glucose level was analyzed calorimetric procedures kits developed by Diamond  Diagnostics kits Cairo, Egypt using 550 n/m. According to (Trinder, 2000).

Determination of triglycerides

Fully enzymatic determination of total triglycerides in serum was estimated spectrophtomtrically at 500ηm according to the method of (Wahlefeld, 1974) of the enzymatic hydrolysis of triglycerides using Stanbio kits followed by determination of the liberated glycerol by colorimetry.

Determination of serum total cholesterol

Enzymatic determination of cholesterol was carried out according to the method of (Allian et al., 1974) using kits purchased from stanbio (Taxas, USA).

Determination of High Density Lipoprotein (HDL-c) cholesterol

The kits were provided from Stanbio, Lab., Inc. Texas. According to (Warnick et al., 1983). Low density Lipoprotein (LDL-c) cholesterol is precipitated from serum by magnesium chloride/ dextran sulfate reagent.

High density lipoprotein (HDL-c) cholesterol is then determined in supernatant using cholesterol reagent.

Determination of low Density lipoprotein (LDL-c) cholesterol

LDL was calculated by the difference between total cholesterol, HDL cholesterol and triglyceride according to as the method of (Friedewald et al., 1998) and was calculated using the following equation.

Calculation:

Calculation of Very Low Density Lipoproteins (VLDL-c) cholesterol

VLDL-c could be estimated reasonably well as TG concentrations divided by constant (5 for mg/dl), according to (Sniderman et al., 2003; Bairaktari et al., 2005 and Gazi et al., 2006) as follow

Calculation:

Statistical analysis

Data was analyzed with analysis of variance (ANOVA) procedures using the MSTAT-C Statistical software package (Michigan State University, 1983).Where the F-test showed significant differences among means (Duncan)  multiple range test (1955) was performed at the 0.05 level of probability to separate means.

Results and Discussion

Table (2): Effect of different concentrations of cinnamon and ginger on blood serum glucose levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter within the same column were not significantly different          

F-test group (G) = (A) 178.6^**  P<0.01                          F-test weeks (W) = (B) 23.6^** P <0.01

F-test (G×W) = (AB) 32.9^**  P <0.01

Effect of different concentrations of cinnamon and ginger on blood serum glucose levels (mg/dl) in the different diabetic rat groups

The results given in the  table (2) revealed that blood serum glucose levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups at (p<0.01).

The results showed that the mean blood serum glucose levels in diabetic rats groups treated with either cinnamon or ginger (3, 4, 5, 6, 7 and 8) recording (233.73, 214.42, 220.28, 200.28, 200.75 and 187.13 mg/dl); respectively were significantly decreased as compared to diabetic group (2) fed on basal diet without any treatment recording 304.45 mg/dl. however, these levels were significantly increased than that of the control group of rats (103.23mg/dl) while, group (9) standard fed on basal diet plus (Atorvastatin 10mg + Gliclagide 30mg) was recorded 137.43 mg/dl by the end of the feeding time of experiments (the 6^th week). These results of this present were in accordance with those reported by (Kamble and Rambhimaiah  2013), (Naserzadeh et al., 2014) and (Muhamed et al., 2016) in cinnamon while, the results content in ginger was agreement (Elshater et al., 2009), (Abdulrazaq et al., 2010), (Jafri et al., (2010), (Diaa and Al-Azhary  2011), (Manal and Anfenan 2014), (Khandouzia et al., 2015), (Hossain and Islam 2016).

These findings were in agreement with (Khan et al., 1990; Shalaby and Samar 2010 and Al-Jamal and Rasheed, 2010). hypoglycemic effect of cinnamon could be due to Insulin Potentiating Factor (IPF) that was isolated from cinnamon, which increased the activity of insulin 3 folds in glucose metabolism in the epididymal fat cells of rat. Later, this isolated factor (IPF) from cinnamon was termed as Methyl Hydroxy Chalcone Polymers (MHCP). They reported that MHCP was increased insulin dependent glucose metabolism many folds (more than 3 folds) when it was examined during in vitro studies (Broadhurst, et al., 2000). Others explained that MHCP caused fat cells more responsive to insulin through activating the insulin-receptor-kinase enzyme that causes binding of insulin to the cells and inhibiting insulin-receptorphosphatase enzyme that blocks this binding process thereby MHCP leads to maximal phosphorylation of the insulin receptor, which is associated with increased insulin sensitivity and enhances glycogen synthase activity (Khan et al., 2003; Anderson et al., 2006; Kim et al., 2006 and Nadia et al., 2011).(Ugochukw and Babady, 2003) mentioned that cinnamon increases the production of Glucose-6-Phosphate Dehydrogenase (G-6-PDH) in the liver in which G-6-PDH leads to reduce glucose transporting by pentose phosphate pathway (pentose shunt) and storage of glucose as a glycogen in the liver. Cinnamon was mentioned in other study to activate the function glucokinase enzyme which in turn stimulates Glucose Transporter-4 (GLUT4) for entering of glucose to the hepatic cells (glycogenesis) and adipocytes lead to increase in glycogen storage available for energy production.

 (Ramudu et al., 2011 and Chakraborty et al., 2012) revealed that ginger has been shown to modulate insulin release in rat pancreatic β-cells, thus enhanced plasma insulin levels in conjunction with lowered blood glucose, this may be due to 6-gingerol, which is considered an active component in ginger, so showed a protective effect on pancreatic β-cells and restored the plasma insulin level.

(White, 2007) reported that ginger is contain over 20 phenolic compounds, these compound displays diverse biological activities such as antioxidant, antidiabetic, hypoglycemic and a lose reductase inhibitory properties.

Kar et al., (1999) reported that, the inorganic part of a medicinal plant contains mainly mineral elements, which are responsible for the hypoglycemic activity. In support of this view, a number of essential minerals (Ca, Zn, K, Mn and Cr), are known to be associated with the mechanisms of insulin release and its activity in different animals and in human beings (Castro, 1998).

Table (3): Effect of different concentrations of cinnamon and ginger on serum triglycerides levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter with in the same column were not significantly different

F-test group (G) = (A) 112.6** P<0.01                          F-test weeks (W) = (B) 33.25** P<0.01

F-test (G×W) = (AB) 5.09**  P<0.01

Effect of different concentrations of cinnamon and ginger on serum triglycerides levels (mg/dl) in the different diabetic rat groups

The results given in the table (3) revealed that serum triglycerides levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups at (p<0.01).

The results of the study showed that the mean triglyceride levels in diabetic rats groups treated with either cinnamon and ginger (3,4,5,6,7 and 8) recording (212.1, 197.4, 200.4, 186.9, 174.8 and 154.2 mg/dl); respectively were significantly decreased as compared to diabetic group (2) fed on basal diet without any treatment recording 246.8mg/dl. However, these levels were significantly increased than that of the control group of rats (108.7 mg/dl). While, group (9)standard fed on basal diet plus (Atorvastatin 10 mg + Gliclagide 30 mg)was recorded 131.2 mg/dl by the end of the feeding time of experiments (the 6^th week).

These results are in agreement with (Gullapalli et al., (2013), (Khogali et al., 2014) and (Amal et al., 2016)in cinnamon while, the results content in ginger was agreement with (Ramudu et al., 2011), (Hussein, 2012), (Maisa et al., 2016) and (El-Sayed and Reham 2016).

Table (4): Effect of different concentrations of cinnamon and ginger on serum total cholesterol levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD 

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter with in the same column were not significantly different

F-test group (G) = (A) 106.7** P<0.01                          F-test weeks (W) = (B) 80.45** P<0.01

F-test (G×W) = (AB) 11.98** P<0.01

Effect of different concentrations of cinnamon and ginger on serum total cholesterol levels (mg/dl) in the different diabetic rat groups

The results given in the table (4) revealed that total cholesterol levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups and feeding weeks at (p<0.01).

The results showed that the mean total cholesterol levels in diabetic rats groups treated with either cinnamon or ginger (3, 4, 5, 6, 7 and 8) recording (220.0, 211.7, 214.4, 201.2, 200.7 and 186.6 mg/dl); respectively were  significantly decreased as compared to diabetic group (2) fed on basal diet without any treatment recording 255.6 mg/dl. However, these levels were significantly increased than that of the control group of rats (145.4 mg/dl). While, group (9) standard fed on basal diet plus (Atorvastatin 10mg + Gliclagide 30mg)was recorded 146.8 mg/dl by the end of the feeding time of experiments (the 6^th week). These results are in agreement with (Srinivasan and Sambaiah1991) reported that feeding rats with ginger significantly elevated the activity of hepatic cholesterol 7-alpha-hydroxylase which is a rate-limiting enzyme in the biosynthesis of the bile acids and stimulates the conversion of cholesterol to bile acids  leading to the excretion of cholesterol from the body. In support of this view, the study of (Bhandari et al., 1998^b) revealed that posttreatment with ginger extract to the cholesterol-fed rabbits for 70 days resulted in less marked hyperlipidaemic when compared to the pathogenic rats. Moreover, the marked hyperlipidemia that characterizes the diabetic state may therefore be regarded because of the uninhibited actions of lipolytic hormones on the fat depots due to the absence of insulin (Goodman and Gilman1985).

(Thomson et al., 2002 and Verma et al., 2004) stated that ginger significantly lowered serum total cholesterol, LDL, VLDL and triglycerides and raised HDL., their results to the reducing effect of ginger acted on cholesterol biosynthesis in the liver and may stimulate cholesterol's conversion to bile acid and increase its fecal excretion.

Table (5): Effect of different concentrations of cinnamon and ginger on the serum High Density Lipoprotein (HDL) levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter with in the same column were not significantly different.

F-test group (G) = (A) 156.4** P<0.01                          F-test weeks (W) = (B) 55.35** P<0.01

F-test (G×W) = (AB) 6.343** P<0.01

Effect of different concentrations of cinnamon and ginger on the serum High Density Lipoprotein (HDL) levels (mg/dl) in the different diabetic rat groups

The results given in the table (5) revealed that High Density Lipoprotein (HDL) levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups and feeding weeks at (p<0.01).

The results showed that the mean HDL levels in diabetic rats groups treated with either cinnamon or ginger (3, 4, 5, 6, 7 and 8) recording (26.03, 28.80, 29.00, 30.72, 30.50 and 32.02 mg/dl); respectively were significantly increased as compared to diabetic group (2) fed on basal diet without any treatment recording 21.39 mg/dl. However, these levels were significantly decreased than that of the control group of rats (39.33 mg/dl). While, group (9) standard fed on basal diet plus (Atorvastatin 10 mg + Gliclagide 30 mg)was recorded 33.50mg/dl by the end of the feeding time of experiments (the 6^th week).

These data are in agreement with (Hussein, 2012) reported that the effect of daily intraperitoneal injection of ginger extract for four weeks on some biochemical parameters (serum cholesterol, HDL, LDL) and kidney functions (urea, uric acid, creatinine), were significant decrease (p<0.01), while (HDL) appeared significant increase when compared with control group antioxidative properties of ginger represent the main way for protecting vascular wall against oxidative stress induced in diabetic cholesterol-fed animals. This effect may work independently of ginger ability to low increased lipid parameters associated with diabetic complications. While, an increased oxidative stress has been shown to be increased in both insulin dependent diabetes.

Table (6): Effect of different concentrations of cinnamon and ginger on the Low Density Lipoprotein (LDL) Levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter with in the same column were not significantly different

F-test group (G) = (A) 178.5** P<0.01                          F-test weeks (W) = (B) 5.231** P<0.01

F-test (G×W) = (AB) 11.36** P<0.01

Effect of different concentrations of cinnamon and ginger on the Low Density Lipoprotein (LDL) Levels (mg/dl) in the different diabetic rat groups

The results given in the table (6) revealed that Low Density Lipoprotein (LDL) levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups at (p<0.01).

The results of the study showed that the mean LDL levels in diabetic rats groups treated with either cinnamon or ginger (3, 4, 5, 6, 7 and 8) recording (150.13, 142.40, 143.22, 131.57, 137.95, and 125.75 mg/dl); respectively were significantly decreased as compared with diabetic groups (2) fed on basal diet without any treatment recording 178.27 mg/dl. However, these levels were significantly increased than that of the control group of rats (81.20 mg/dl). While, group (9) standard fed on basal diet plus (Atorvastatin 10mg + Gliclagide 30mg)was recorded 91.17 mg/dl by the end of the feeding time of experiments (the 6^th week).

Ginger lowered the level of total cholesterol, TG, VLDL and LDL and improved the level of hyperlipidemic and diabetic rats these results were in agreement with those previously reported (Fuhrman et al., 2000 and Nicoll and Henein  2009) found that The adding ginger rhizome powder to the food could be useful in the management of cardiovascular disease in which atherosclerosis is the most important factor may be due to the presence poly-phenolic and flavonoids may prevent coronary artery disease by reducing plasma cholesterol level or by inhibiting LDL oxidation, a process which is through to play a key role in the pathogenesis of atherosclerosis.

Table (7): Effect of different concentrations of cinnamon and ginger on the Very Low Density Lipoprotein (VLDL) levels (mg/dl) in the different diabetic rat groups.

Values are presented as mean ± SD

For each group of rats n = 10                                          S.E. Standard Error

Values followed by the same letter with in the same column were not significantly different

F-test group (G) = (A) 231.5** P<0.01                          F-test weeks (W) = (B) 63.5** P<0.01

F-test (G×W) = (AB) 7.12** P<0.01

Effect of different concentrations of cinnamon and ginger on the Very Low Density Lipoprotein (VLDL) levels (mg/dl) in the different diabetic rat groups

The results given in the table (7) revealed that Very Low Density Lipoprotein (VLDL) levels showed significant differences among all the nine studied groups, all feeding weeks as well as interaction between studied groups at (p<0.01).

The results of the study showed that the mean (VLDL) levels in diabetic rats groups treated with either cinnamon and ginger (3, 4, 5, 6, 7 and 8) recording (41.38, 38.68, 40.90, 37.07, 30.17 and 26.67 mg/dl); respectively were significantly decreased as compared to diabetic group (2) fed on basal diet without any treatment recording 49.20mg/dl. However, these levels were significantly increased than that of the control group of rats (22.05 mg/dl). While, group (9) standard fed on basal diet plus (Atorvastatin 10 mg + Gliclagide 30 mg)was recorded 25.07 mg/dl by the end of the feeding time of experiments (the 6^th week).

Cinnamon lowered the level of total cholesterol, TG, VLDL and LDL and improved the level of hyperlipidemic and diabetic rats. The observation of higher level of serum lipids in diabetic rats in many studies were found.

(Brahmachari, et al., 2009) mentioned that major compounds of cinnamon (cinnamaldehyde, cinnamyl acetate and cinnamyl alcohol) are converted into cinnamic acid by oxidation and hydrolysis, respectively. In the liver, cinnamic acid oxidized to benzoate that exists as sodium benzoate or benzoyl-CoA. Sodium benzoate was found to reduce the level of cholesterol in vivo in mice.

Conclusion:

In conclusion on the basis of above mentioned data. There were significant differences between untreated group and all other (8) studied groups of the experimental rats (p < 0.01) in the serum glucose, serum triglycerides, total cholesterol, HDL cholesterol, LDL cholesterol and VLDL cholesterol. The best improvement on the serum glucose, triglycerides and cholesterol fractions in diabetic standard group was fed on basal diet plus (10 mg Atorvastatin + 30 mg Gliclagide) followed by diabetic group was fed on basal diet plus 10% of both cinnamon and ginger then diabetic group was fed on basal diet plus 10% cinnamon and diabetic group was fed on basal diet plus 10% ginger.