Journal of Genetics and Molecular Biology

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Research Article - Journal of Genetics and Molecular Biology (2021) Volume 5, Issue 3

In vitro anti oxidant activity of kaempferol micro emulsion.

Minal S Patil1*, V R Patil2

1Department of DCSs, Annasaheb Ramesh Ajmera college of Pharmacy, Nagaon, Dhule, Maharashtra, India

2Department of TVESs, Hon’ble L. M. C. College of Pharmacy, Faizpur, Jalgaon, Maharashtra, India

*Corresponding Author:
Dr. Minal S Patil
Department of DCSs
Annasaheb Ramesh Ajmera college of Pharmacy
Dhule, Maharashtra, India
E-mail: [email protected]

Accepted date: April 14, 2021

Citation: Patel MS, Patel VR. In vitro anti oxidant activity of kaempferol micro emulsion. J Genet Mol Biol 2021;5(3):1-3.

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Abstract

Flavonoids are promising antioxidants. Kaempferol is a natural flavonoid with potent antioxidant activity, but its use is limited because of its low aqueous solubility. The present study sought to investigate the capacity of single herbal formulations of Kaempferol to act as nitric oxide radical (NO), 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical antagonists using in vitro models. Observed Results indicates that optimised batch of micro emulsion exhibited potent antioxidant activity 

Keywords

Kaempferol Micro emulsion, DPPH, Ascorbic acid, Nitric oxide, Sodium nitro prusside etc.

Introduction

Nature is always a source of medicinal agents for thousands of years and an impressive number of modern drugs are isolated from natural sources, many based on their use in traditional medicine [1]. Antioxidants are compounds which have ability to bring either delay or inhibit the oxidation processes which occur under the influence of atmospheric oxygen or reactive oxygen species. Antioxidants are involved in the defense mechanism of the organism against the pathologies associated to the attack of free radicals [2].

Flavonoids, plant polyphenols are a group of plant secondary metabolites characterized by a diphenyl propane structure. They are widely distributed in the plant kingdom and are common constituents of fruits, vegetables [3]. Many of them have been used as traditional medicine in India and other Asian countries for more than thousands of years.

These plant polyphenols are with strong antioxidant capacities and thus largely contribute to the pharmaceutical and dietary properties of plant derived food. Antioxidant compounds have been receiving great attention from natural products consumers and researchers due several pharmacological properties. Antioxidants played an important role in lowering oxidative stresses caused by reactive oxygen species. Antioxidants are substances that employ various mechanisms to scavenge free radicals by inhibiting their formation or interrupting their propagation [4]. Thus, through various mechanisms antioxidants have the ability to inhibit the adverse effects of oxidative stress. The Kaempferol, a yellow compound with a low molecular weight and molecular formula is a common natural flavonoid which representative of the subcategory of flavonol that commonly found in many plant-derived foods and in plants used in traditional medicine also reported its various therapeutic effects like anticancer, antioxidant, antiinflammatory and hepatoprotective. However, the oral bioavailability of kaempferol is relatively low because of its low lipid solubility and its limited membrane permeability. Kaempferol has been identified in many botanical families and has been found in Pteridophyta, Pinophyta and Magnoliophyta. Kaempferol revealed low to moderate absorption, which results poor bioavailability ~2%.It is hydrophobic in nature and freely soluble in methanol [5].

Materials and Methods

Chemicals and reagents

DPPH, sample/s stock, Ascorbic acid, Sodium nitroprusside, Sulphanilamide, Potassium ferricyanide, Ferric chloride, N-(1- naphthyl) ethylenediamine dihydrochloride. All other reagents were of analytical grade [6].

Kaempferol collection

Kaempferol sample was purchased from Yucca Laboratories Pvt. Ltd, Mumbai.

DPPH radical scavenging activity

DPPH [1, 1-Diphenyl-2-picryl hydrazyl] is a stable free radical, which shows absorbance at 517 nm. The antioxidant reacts with DPPH and converts it to 1, 1-Diphenyl-2-picryl hydrazine which do not absorb at 517 nm [7]. The antioxidant activity of the micro emulsion of kaempferol was determined in terms of hydrogen donating or radical scavenging ability using the stable radical DPPH carried out by using the method of Molyneux. To 1 ml of DPPH solution, equal amount of test compound at various concentrations (20-100 ug/ml) were added in a final volume of 2.0 ml [8]. After incubation for 20 minutes at room temperature, absorbance due to changes in color from deep violet to light yellow were recorded at 517 nm. The control solution was prepared by mixing ethanol (3.5 mL) and DPPH radical solution (0.3 mL). The different concentrations of ascorbic acid were used as reference compound. Lower absorbance of the reaction mixture indicated higher free radical activity. Radical scavenging activity was expressed as the inhibition percentage of free radical by the sample [9].

Nitric oxide radical scavenging activity

When sodium nitroprusside was mixed with aqueous solution at physiological pH, suddenly it generates nitric oxide, which reacts with oxygen to produce nitrite ions. Nitric oxide scavengers compete with oxygen leading to reduced production of nitrite ions [10].

Nitric oxide radical scavenging activity was measured spectrophotometrically according to the method described by Govindharajan. About 1 ml of Sodium nitroprusside (5 mM) in phosphate buffer was mixed with different concentrations of the samples in phosphate buffer. [11].

The tubes were then incubated at 25°C for 2 h. After incubation 1.5 ml of reaction mixture was removed and diluted with 1.5 ml of Greiss reagent [1% sulphanilamide, 2% Ophosphoric acid and 0.1% of N-(1-naphthyl) ethylenediamine dihydrochloride]. The absorbance was measured spectrophotometrically at 546 nm. [12].

Control tube was maintained with all chemicals excluding sample. Radical scavenging activity was expressed as the inhibition percentage of free radical by the sample and was calculated.

Results and Discussion

Results of present study showed that all kaempferol samples had significant levels of radical scavenging activity in a dose dependent manner (Table 1 and Figure 1).

Concentration
(ug/ml)
Percentage Inhibition (Mean ± SEM) (n=3)
Sample Standard
20 11.60 ± 0.033 21.14 ± 0.066
40 18.43 ± 0.147 24.62 ± 0.066
60 22.45 ± 0.140 29.17 ± 0.033
80 28.27 ± 0.066 33.69 ± 0.033
100 32.59 ± 0.091 39.39 ± 0.066
IC50 49.64 49.55

Table 1. DPPH free radical scavenging activity of optimised batch of microemulsion containing kaempferol.

european

Figure 1: DPPH free radical scavenging activity of optimised batch of microemulsion containing kaempferol.

The DPPH assay is purely based on the assumption that an antioxidant serves as a hydrogen donor and thus reduces the DPPH free radicals (the color turns from deep violet to light yellow) [13]. In this study of Nitric oxide radical scavenging activity results showed that all samples had significant levels of radical scavenging activity in a dose dependent manner (Table 2 and Figure 2).

Concentration
(ug/ml)
Percentage Inhibition (Mean ± SEM) (n=3)
Sample Standard
20 20.43 ± 0.036 30.54 ± 0.070
40 26.52 ± 0.118 38.97 ± 0.033
60 33.73 ± 0.066 42.68 ± 0.170
80 37.61 ± 0.066 47.75 ± 0.115
100 42.18 ± 0.066 50.68 ± 0.170
IC50 49.51 49.38

Table 2. Nitric oxide radical scavenging activity of optimised batch of micro emulsion containing kaempferol.

european

Figure 2: Nitric oxide radical scavenging activity of optimised batch of microemulsion containing kaempferol.

DPPH Radical Scavenging Activity.

Conclusion

The results obtained from the study confirmed the benefits of the medicinal plants. In fact, some flavonoids present in them demonstrated high antioxidant activities and also a low acute toxicity effect. These data show that the optimized batch of micro emulsion had an antioxidant activity more than 40% at100 μg/ml in Nitric oxide radical scavenging activity and more than 30% at100 μg/ml in DPPH free radical scavenging activity [14]. This confirms that flavonoids are non negligible compounds [15].

References

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