Identification and Quantification of Flavonoids in Musa Paradisiaca by Using Colorimetric Method

V. Shirisha, K. Neha Subadra Devi, K. Gayathri Susmitha, Y. Ravi Teja, CH. Shainy, M. Bapi Raju,

doi:10.5281/zenodo.15121449

Research Paper | Open Access
Volume 03 | Issue 04 | Article Id IJPS/250303393

Identification and Quantification of Flavonoids in Musa Paradisiaca by Using Colorimetric Method
V. Shirisha* K. Neha Subadra Devi K. Gayathri Susmitha Y. Ravi Teja CH. Shainy M. Bapi Raju
A.K.R.G. College of Pharmacy, Affiliated to JNTUK, Nallajerla, East Godavari, Andhra Pradesh-534112.

Abstract

Musa paradisiaca, a widely distributed medicinal plant, is known for its diverse pharmacological properties, including antimicrobial, anti-cancer, anti-oxidant, anti-diabetic, anti-inflammatory, and wound healing effects. The presence of bioactive flavonoids contributes significantly to its therapeutic potential. This study aims to determine and quantify the flavonoids content in musa paradisiaca leaves using a colorimetric method. Flavonoids were extracted using soxhlet extraction techniques, and their concentration was measured colorimetrically. The absorbance was recorded at a specific wavelength of 450nm, and a calibration curve was established using a standard rutin. The results indicated 6.2% of flavonoids in the leaf extract, demonstrating the potential of Musa paradisiaca as a source of bioactive compounds. This study provides a simple, cost effective, and reliable method for flavonoids quantification, which could be useful for further pharmacological studies.

Keywords

Quantification, Flavonoids, Colorimetric Method, colorimetrically, anti-inflammatory.

Introduction

A colorimeter is a device that is used in Colorimetry. It refers to a device which helps specific solutions to absorb a particular wavelength of light. The colorimeter is usually used to measure the concentration of a known solute in a given solution with the help of the Beer-Lambert law.

The colorimeter was invented in the year 1870 by Louis J Duboscq.

Principle of Colorimeter

It is a photometric technique which states that when a beam of incident light of intensity Io passes through a solution, the following occur:

- - A part of it is reflected which is denoted as Ir
  - A part of it is absorbed which is denoted as Ia
  - Rest of the light is transmitted and is denoted as It

Therefore, Io = Ir + Ia + ItTo determine Ia the measurement of Io and It is sufficient therefore, Ir is eliminated. The amount of light reflected is kept constant to measure Io and It.

Advantages of Colorimeter:

Simple and Easy to Use - Requires minimal training and is straightforward in operation.
Quick Results - Provides rapid and reliable measurements.
Cost-Effective - More affordable compared to spectrophotometers.

Disadvantages of Colorimeter:

Limited to Colored Solutions - Cannot analyze colorless solutions or gases effectively.
Less Precise Than Spectrophotometers - Not suitable for highly accurate broad-spectrum analysis.
Affects by Turbidity and Impurities - Suspended particles can interfere with the accuracy of results.

MATERIALS:

Musa paradisiaca leaves, Ethanol, Rutin, Sodium nitrite, Aluminium chloride, Sodium hydroxide, Petroleum gelly, Distilled water.

Apparatus:

Soxhlet apparatus, RBF, Condenser, Whatman filter paper, Spatula, Funnel, Tripod stand, Measuring cylinder, Filter paper, Beaker, Volumetric flask (10ml,100ml), Glass rod, Pipette,Ttest tubes, Cuvettes.

Instruments:

Digital balance, Colorimeter, Heating mantle.

Plant Collection:

The collected fresh leaves of Musa paradisiaca were washed and dried in shade. After drying plant leaves were finely powdered and kept in a well closed container. For further extraction procedure.

Method Of Extraction:

Soxhlet method:30 grams of finely powdered Musa paradisiaca leaves are packed in a thimble and placed in the sample holder. And pour 350 ml of ethanol into it. Extraction was carried out for 12 hr at 80°C. Later on, the extract was filtered, and the filtrate was stored for further use.

Phytochemical Screening

Identification Test for Alkaloids:

Test	Procedure	Observation
1.Dragendoff Test	Take 2 ml extract and add 2ml of draendorff reagent to it.	A orange red precipitate was formed. It indicates the presence Alkaloids.
2.Hager’s Test	Take 2 ml extract and add 2 drops of aqueous picric acid solution.	Acreamy white precipitate was not formed. It indicates the absence of alkaloids.

Dragendorff’s test

Identification Tests for Flavanoids:

Test

Procedure

Observation

1.Ferric chloride test

Take 2ml extract and add Ferric chloride solution to it.

Greenish black colour was formed which indicates

the presence of Flavonoids.

2.Alkaline reagent test

Add few drops of sodium hydroxide to a test solution.

Yellow colour was formed which indicates the absence of flavonoids.

Ferric chloride test

Identification Test for Carbohydreates:

Dragendorff’s test

Identification Tests for Flavanoids:

Test

Procedure

Observation

1.Ferric chloride test

Take 2ml extract and add Ferric chloride solution to it.

Greenish black colour was formed which indicates

the presence of Flavonoids.

2.Alkaline reagent test

Add few drops of sodium hydroxide to a test solution.

Yellow colour was formed which indicates the absence of flavonoids.

Ferric Chloride Test

Identification Test for Carbohydreates:

Test	Procedure	Observation
1.Molisch’s test	Take 2ml extract and add αnaphthol and few drops of concentrated H2SO4.	A purple ring was formed at the point of contact between the acidic and which indicates the presence of carbohydrates.
2.Fehling’s test	Take 2ml of extract and add equal quantities of Fehling’s A &B solutions.	A brick red precipitate was formed which indicates the presence of carbohydrates.

Identification Test for Terpinoids

Salkowski Test

Test	Procedure	Observation
1.Salkowski test	Take 5ml extract and add 2ml chloroform and evaporate on water bath then add 3ml concentrated H2SO4.	Reddish brown interface was formed which indicates the presence of terpenoids.

Method Of Preparation:

Preparation of Rutin solution:

Accuratelyweighed1gmofRutinandaddto100mlvolumetricflask.
Added30mlofethanolintoittodissolveandmakeupthevolumeto 100ml.

Preparation of sodium hydroxide solution:

Accuratelyweighed4gmsofNaoHandaddto100mlvolumetricflask.
Added30mlofdistilledwaterintoittodissolveandmakeup thevolumeto 100ml.

Preparation of sodiumnitrite:

Accuratelyweighed5gmofsodiumnitriteandaddto100mlvolumetricflask.
Added30mlofdistilledwaterintoittodissolveandmakeupthevolumeto100ml.

Preparation of Aluminiumchloride:

Accuratelyweighed10gmofAlcl3andaddto100mlvolumetricflask.
Added30mlofdistilledwaterintoittodissolveandmakeupthevolumeto100m.

Selection Of Wavelength:

For1mlofRutinsolution10mlvolumetricflask.
Makeupthevolumeto10ml.
Measuretheabsorbanceoftheabovesolutionatdifferentwavelenghths.(450nm,470nm,510nm,520nm,540nm,570nm,600nmand670nm) against blank.

Themaximumabsorbanceisat450nm-λmaxis450nm

S. No	Wavelength (Nm)	Concentration (Mg/Ml)	Absorbance
1.	450nm	1mg/ml	0.47
2.	470nm	1mg/ml	0.41
3.	510nm	1mg/ml	0.21
4.	520nm	1mg/ml	0.09
5.	540nm	1mg/ml	0.04
6.	570nm	1mg/ml	0.22
7.	600nm	1mg/ml	0.02
8.	670nm	1mg/ml	0.01

Preparation Of Calibration Curve:

Five different concentrations (0.2 mg , 0.4mg , 0.6 mg , 0.8 mg , and 1.0 mg) of rutin were taken from standard 1 mg/ml Rutin in five different 10 ml volumetric flasks.
Added 0.3ml of 5% NaNo2 and 0.3ml of 10% Alcl3 and 2ml of 0.4 % NaoH solution.
Then keep it for 6min.
Make up the volume with distilled water upto the mark.
The solution was keep it for 15 mins at room temperature.
The absorbance was measured at 450nm.

S. No	Wavelength (Nm)	Concentration (Mg)	Absorbance
1.	450nm	0.2	0.18
2.	450nm	0.4	0.31
3.	450nm	0.6	0.47
4.	450nm	0.8	0.56
5.	450nm	1.0	0.73

Procedure For Determination of Extract

0.2ml of plant extract was taken in a10ml volumetric flask
Added 0.3ml of 5% NaNo2 and 0.3ml of 10% Alcl3 and 2ml of 4% NaoH solution.
Then keep it for 6min.
Makeup the volume with distilled water upto the mark.
The solution was keep it for 15 mins at room temperature.
Then the absorbance was measured at 450nm.

RESULT AND DISCUSSION

Calculation

S. No	Wavelength (Nm)	Concentration (Mg)	Absorbance
1.	450nm	0.2	0.18
2.	450nm	0.4	0.31
3.	450nm	0.6	0.47
4.	450nm	0.8	0.56
5.	450nm	1.0	0.73

FORMULA:

Concentration of flavonoid content in 30gms (450ml)

Concentration found × Volume of final extract

Total flavonoid content = Quantity of extract taken

= 0.83 ×450

0.2

1867.5mgofrutinin30gmsofpowder.

Concentration in 30gms

% of flavonoids in 30gms = × 100 Weight of powder = 1.8675

30 = 6.225%

The total flavonoid content in Musaparadisiaca leaves was found to be 6.225%

Summary

This study focuses on the identification and quantifiacation of flavonoids content in Musa paradisiaca leaves using a colorimeter method.Flavonoids, known for their medicinal properties, Were extracted using in Soxhlet extraction technique.The quantification was performed by forming a colored complex and the absorbance was measured colorimetrically at 450nm.A standard calibration curve using known flavonoids(rutin) concentrations was used to determine the flavonoids content in the plant extract.The results confirmed 6.225%of flavonoids in Musa paradisiaca leaves,supporting it’s potential for pharmaceutical applications.This method provides a simple,cost effective, and reliable approach for flavonoids analysis,beneficial for quality control and further pharmacological research.

CONCLUSION

The colorimetric method used in this study successfully determined and quantified the flavonoid content in Musaparadisiaca leaves.The extraction and complexation provided are liable and cost-effective approach for Flavonoids analysis .The results confirmed a significant presence of flavonoids, highlighting the medicinal potential of Musaparadisiaca . This method can serveas a valuable tool for quality control in herbal formulations and further pharmacological research.Further studies can focus on optimizing the method for higher sensitivity and exploring the biological activities of the quantified Flavonoid’s.

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