Analysing The Hypolipidemic Activities of The Tea Extracts of Moringa oleifera, Glycyrrhiza glabra and Their Blend at Different Concentrations, Orally Induced on Adult Male Wistar Rats

Herbal teas of Moringa oleifera (Moringa) and Glycyrrhiza glabra (Licorice) were known to have many beneficial effects, their possible hypolipidemic effects were investigated in this paper. Albino wistar rats were fed with aqueous extracts of M. oleifera, G. glabra and their blend teas (10, 30 and 50 mg/kg.BW/ml). The teas were found to lower the serum cholesterol, triglycerides, VLDL, LDL, and atherogenic index at their higher concentration, but were found to slightly increase the HDL as compared to the corresponding normal healthy rats fed with clean water (control). The phytochemicals screening of the teas aqueous extract were also investigated, the chemicals (tannins, saponins, flavonoids, terpenoids and glycosides) which were known to exhibit cholesterol lowering effect by inhibiting its absorption and simultaneous increase its excretion were found to be present in the teas extract. Thus, the study demonstrates and validates that M. oleifera, G. glabra and their blend possess hypolipidemic effect at higher concentration.


Total Flavonoid content
The total flavonoid content of the extracts was determined using a slightly modified method reported by Chung et al., [5]. Briefly, 0.5mL of enzyme digested sample was mixed with 0.5mL methanol, 50μl of 10% AlCl 3 , 50μl of 1mol L -1 potassium acetate and 1.4mL water, and allowed to incubate at room temperature for 30 min. Thereafter, the absorbance of each reaction mixture was subsequently measured at 415 nm. The total flavonoid was calculated using quercetin as standard by making use of a seven point standard curve (0, 20, 40, 60, 80,100 μg/mL). The total flavonoids content of samples was determined in triplicates and the results were expressed as mg quercetin equivalent per gram of the sample.

Total Phenolic content
The total phenolic content of the samples extract was determined by the Folin-Ciocalteu assay as described by Chanda et al [4]. 500μl of Folin reagent was added and mixed with a solution containing 100μL of the extract and 2mL of distilled water. 1.5mL of 7.5% sodium carbonate was then added to the solution and the volume was made up to 10mL with distilled water. The mixture was left to stand for 2 h after addition of the sodium carbonate. The absorbance of the mixture was measured at 760 nm using a Lambda EZ150 spectrophotometer (Perkin Elmer, USA). The standard used was tannic acid and the results were expressed as mg tannic acid equivalents per gram of the sample.

Biochemical assays for lipids
Cholesterol, HDL and triglyceride levels were estimated from serum by using a commercial kit package (Randox Laboratories Limited). We used standard commercial kits for analysis as recommended by the manufacturer of these kits. LDL and VLDL-cholesterol were calculated following the method by Johnson et al. [12], while the atherogenic index was calculated by using the method described by Muruganandan et al. [17]. LDL = TC -(HDL + Triglyceride/5) VLDL = TC-HDL-LDL Atherogenic index = (TC-HDL)/HDL

Statistical analysis
Results are expressed as mean±SEM (standard error mean) and subjected to one-way analysis of variance (ANOVA) followed by Dunnett's test and values with p<0.05 were considered to be statistically different.

Results and Discussions
Phytochemical investigation was performed and the following compounds were identified in the teas extracts as shown in Table 1. The phenolic and flavonoid contents of Moringa, Licorice and their blend vary as shown in figure 1 and 2. Plant phenolics constitute one of the major groups of compounds acting as primary antioxidants or free radical terminators, it was reasonable to determine their total amount in the selected plant extracts [31]. Phenolics and polyphenolics (polymeric phenolics) can provide relief from certain physical ailments and degenerative diseases in humans, including the reduction of cardiovascular disease and certain cancers [26,1]. Therefore, it is not surprising that the extraction and analysis of phenolics from plants and other food sources have been extensively studied [18,6].

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IJPPE Volume 3     Flavonoids present in food of plant origin are also potential antioxidants [24]. Moringa, Licorice and their blend are good source of flavonoid (Figure 1). The antioxidative properties of flavonoids are due to several different mechanisms, such as scavenging of free radicals, chelation of metal ions, such as iron and copper, and inhibition of enzymes responsible for free radical generation [2].
Elevated level of blood cholesterol especially LDL-C is a known major risk factor for CHD whereas HDL-C is cardio protective. Treatment with aqueous extract of Moringa tea, Licorice tea and their blend, at three different doses, significantly decreased the levels of total cholesterol and LDL-C with respect to the normal control without tea extract [29] (Table 2, 3 and 4). This can be deduced from the results, as the concentration increases from 10mg/kg BW to 50mg/kg BW the values of total cholesterol decreases: for Moringa, from 148.87 ± 1.0 a to 80.70 ± 1.41 c , for Licorice from 139.85 ± 0.71 a to 78.95 ± 3.19 c and for their blend from 152.13 ± 1.06 a to 102.01 ± 3.90 b comparing with the control rats value 69.92 ± 0 .75 a . Likewise LDL-C values decreases with the increase in extracts concentration: for Moringa, it decreases from 78.59 ± 0.36 a to 13.23 ± 0.46 c , for Licorice, it decreases from 60.71 ± 1.19 a to 12.60 ± 3.02 c and for their blend from 75.24 ± 1.17 a to 30.19 ± 2.07 b . Moringa and Licorice show a very significant decreases compare to control (17.33 ± 0.37) reinforcing their individual ability to lower LDL-C. The benefits and therapeutic significance of the two teas are visible in the average values of the blended sample as they exhibit the combination of the individual sample potential cholesterol-reducing ability.
Atherogenic index indicates the deposition of foam cells or plaque or fatty infiltration or lipids in heart, coronaries, aorta, liver and kidneys. The higher the atherogenic index, the higher is the risk of the above organs for oxidative damage [16]. Atherogenic index was significantly reduced as the concentration of M. oleifera (from 2.44 ± .06 a to 0.81 ± .006 c ) and Glycyrrhiza glabra (from 2.13 ± 0.039 a to 0.75 ± 0.056 c ) increases. However, the blended groups (from 2.32 ± 0.006 a to 1.22 ± .053 b ) show a slight decrease in atherogenic index compared to control value (1.06 ± .03 a ).
Rich fatty food causes the oxidative stress which does increases production of reactive oxygen species. A lot of scientific journals provides ample direct or indirect evidence that overproduction of ROS can induce cellular damage via oxidation of critical cellular components such as membrane lipids, proteins, and DNA. Since the result of the study indicated that the aqueous extracts of M. oleifera, Glycyrrhiza glabra and their blend, have beneficial effect on lipid profile, it will be good to identify what may be responsible for reducing potential of the teas.
Plant phytochemicals (such as tannins, glycosides, terpenoids, alkaloids, saponins, and flavonoids etc) inhibit the absorption of dietary cholesterol, but the resulting decrease in serum cholesterol has been slight [25]. Moringa tea has been shown to contain tannins, saponin, glycosides and terpenoid in phytochemical screening. Licorice tea has been shown to possess alkaloids, flavonoids and glycoside while the tea blend contains tannins, saponins, glycosides, flavonoids and terpenoid which are combination of the phytochemicals of the two teas ( Table 1). The cholesterol lowering effect may be due to the inhibition in reabsorption of cholesterol from endogenous sources in association with a simultaneous increase in its excretion.
Conclusively, the observed cholesterol reducing action of the aqueous extract of Moringa oleifera, Glycyrrhiza glabra and their blend which may be responsible by their intrinsic phytochemicals, phenolic and flavonoid contents indicates their hypolipidemic activities.

International Journal of Pharmacology, Phytochemistry and Ethnomedicine
Vol. 3