BIOCHEMICAL STUDIES OF Clitoria ternatea AND ITS USE AS BIOGEL

Abstract

Clitoria ternatea (CT), often known as butterfly pea, is a perennial herbaceous plant that grows around the world, but is most common in tropical areas. It is a member of the Fabaceae family. The plants' genotypes are generally identified by flower colours like blue, white, and pink. The plant has great quality of numerous medicinal properties presented in every part of plant like root, leaf, flower, and seed. The plant that haslong been employed in conventional herbal remedies to treat a wide range of illnesses do so because they contain a large number of bioactive chemicals and antioxidant enzymes. The present research objectives were to evaluate of its phytochemicals and different antioxidant properties through qualitative and quantitative analysis by 60% ethanol extraction. The selection of plant for the research study was the blue flower genotype of root, leaf, flower, and seed and compared its bioactive compound. The formation of biogel from Clitoria ternatea's blue flower component was the final objective of this study. The formulated bio gel was evaluated by its physical evaluation, PH measurement, spreadability, stability, viscosity, skin sensitivity test, toxicity, and inhibitory effect of microbes.

The qualitative properties of C. ternatea were tested by appropriate methods to assess secondary metabolites or bioactive compounds such as total carbohydrates, alkaloids, amino acids and proteins, flavonoids, glycosides, tannins, phenols, saponin, sterols, terpenoids, quinones, oxalates and resins based on deep to light colour present or absent in four different part of the root, leaf flower and the seed of CT plant.

The qualitative test of the above secondary metabolites revealed that the carbohydrates were present in all tissues. In seeds tennins, amino acids, proteins, sterols, and terpenoids were strongly present whereas resins, phenols, flavonoids, glycosides, and alkaloids were more present, while quinones were less present. The oxalates and saponins were absent in the seed. However, the flowers contain highly present alkaloids, flavonoids, phenols, tannins, terpenoids, and quinones but more present resins, and less present oxalates, sterols, amino acids, and proteins, glycosides and found absent of saponin. Flavonoids, amino acids, proteins, and terpenoids were less prevalent in the root section of Clitoria ternatea, but alkaloids,

vii

glycosides, phenols, tannins, sterols, quinones, oxalates, and resins were not present. Although phenols were abundant in the leaves, Clitoria ternatea lacked alkaloids, saponin, and oxalates, and had more flavonoids, tannins, sterols, and quinones. Terpenoids, resins, amino acids, proteins, and glycosides were less prevalent.

The quantitative analysis was estimated through selected parameters such as total carbohydrates, total phenols, flavonoids, tannins, and anthocyanin in different parts (seeds, flowers, leaves, and roots) of Clitoria ternatae. The biochemical study revealed through quantitative analysis that the total carbohydrate was found (89.01 mg Glc/g fresh wt) to be higher in leaf tissue, and the average mean value was obtained 76.75. Similarly, total flavonoids (469.34 Catechine mg/g fresh wt.), the total phenols (93.00 mg GAE/g fresh wt.), total tannins (72.00 mg TAE/g fresh wt.), and total anthocyanins (178.33 mg CGE/g fresh wt.) were presented highest in flower tissue of Clitoria ternetea in comparison to other parts of the plant.

Clitoria ternatea's antioxidant activity was evaluated utilizing the DPPH, FRAP, ABTs, and superoxide radical scavenging tests. The highest DPPH, ABTS, and superoxide activities were observed maximum in flowers i.e. 92.43 mg ascorbic acid/ g FW, 89.36 mgGAE/g FW, and 89 mg ascorbic acid/g FW respectively against standard. Similarly, FRAP activity was maximum in leaves (399 mg BHT/g FW). The observation of DPPH, ABTs, FRAP, and superoxide activities was also recorded in roots, leaves, seeds, and flowers on different concentrations like 0.2, 0.4, 0.6, 0.8, and 1.0 mg/ml through inhibition percentages against standard. The results showed that maximum activity was recorded at 1.0 mg/ml concentration in comparison to other concentrations.

Plant sections of Clitoria ternatea were subjected to an analysis of the antioxidant enzymes superoxide dismutase, peroxidase, polyphenol oxidase, and guaiacol peroxidase. The peroxidase activity was observed highest in flower (391.20 U/ml), followed by leaves (376.33 U/ml), lowest in seed (289.14 U/ml), and the mean value was recorded 340.23 U/ml.

viii

The biogel was formulated with different chemical ingredients with dried CT extract (0.1% and 0.2%) with two formulations against control. It was observed at par in all tests acquired but 0.2% was better than 0.1% formulation of biogel. The results showed that the formulated biogel was deep blue colour, clear, transparent, had good viscosity (2200 cps), good spreadability (25.4 gm. cm/sec), good PH (6-7.04), and homogenous with smooth texture. The skin sensitivity test of biogel was continued for up to seven days regularly, three times a day on the different skin of human beings. It was observed that biogel has no side effects on the skin like irritation, allergy, edema, and rashes after using it. The storage capability of biogel was also very good under various temperatures (20OC, 25OC, and 30OC) and humidity conditions (RH 60%) for duration of three months. The formulated biogel was evaluated through one antifungal (Candida albicans) and three antibacterial (Staphylococcus aureus, Escherichia coli, and Bacillus cereus) activities based on the zone of inhibitions (diameter mm) at three concentrations i.e. 10%, 20% and 30% v/w for 2 to 3 days against control. The highest inhibition zone against positive control obtained from Escherichia coli (14.5 mm/16.5 mm) followed by Staphylococcus aureus (13.2 mm/16.0 mm) and Bacillus cereus (13.1mm/15.1 mm) at 30% concentration of biogel in 42 hours whereas antifungal activity (Candida albicans) was observed (13.3 mm/14.8 mm) at 30% concentration of biogel in 72 hours. The maximum inhibitory zone was seen at a 30% of biogel concentration, as compared to 10% and 20% in specific microbial activity. For the toxicity test biogel of Clitoria ternatea was analyzed through an ICP-MS instrument for the detection of heavy metals like Arsenic and Lead. It was found that 0.01 mg/kg in each could be ignored.

Clitoria ternatea exhibits remarkable antioxidant properties observed in whole plant parts in the present research work. However, flower extract exhibits more antioxidant potential than seed, leaf, and root. All part of plant extract contains many phytochemicals and antioxidant enzymes like Peroxidase, gluaiacol peroxidase, superoxide dismutase, phenolic antioxidant, flavonoids, etc. which is beneficial for disease removal. The overall observation of the present research study concluded that Clitoria ternatea could be used safely for medicated purposes as well as industrial production of herbal medicines.