INTRODUCTION:

Traditional systems of medicines always played important role when it comes to meeting the global health care needs. In India, they are continuing to do so at present and shall play a major role in future as well. The indian system of medicine has unique distinction of having six recognized systems which are Ayurveda, Siddha, Unani and yoga, Naturopathy and Homeopathy.^1,2 Among the six systems of medicine Unani is the type of medicine system which is widely used nowadays The formulations which were used in the project are categorized under unani system of medicine^3,4. The main reason for opting these formulations is that they have good amount of phenols, flavonoids which are responsible for the antioxidant activity of the formulations.

Antioxidants could be used for management of some pathophysiological conditions, which involve free radical, such as cardiovascular and neurodegenerative disease. Flavonoid and phenolic compounds may be useful as antioxidant from natural sources.^5-9

In Unani, Habbe means a pill prepared from the powder of ingredients, some solution, and binder.¹⁰Habb-e-Azaraqi is an Unani medicine having Muharrik (Stimulant), and Muqawwi-e-Asab (nervine tonic) action. This medicine is useful in Nazla Barida Muzmina, falij (paralysis), laqwa (facial palsy), niqrus (gout), Waja-ul-Mufasil (arthritis), Neuralgia, hemiplegia, sciatica etc. All ingredients in Habb -e - Suranjan have anti-inflammatory action and exert anodyne effect.^11-14

To boost our country’s economy from sales and exports of herbal drugs and formulations there is urgent need to adopt newer scientific technologies in proper identification, processing, characterization, ensuring uniformity, pharmacological screening, clinical studies and documentation of research. Such scientific aspects will project herbal products in a proper-perspectives and help in sustaining in global market.^15-19

In the present study, an attempt is made to check the efficacy of few marketed formulations by screening phytoconstituents, total antioxidant capacity and nitric oxide antioxidant capacity using established models.

MATERIALS AND METHODS:

Collection of material:

The Herbal formulations Habb-e-suranjan and Habb-e-azaraqi were collected from the local retail unani stores, Hyderabad, Telangana State, India. All the chemicals, glass ware and equipments used in this study were obtained from the central store house of the institution. The chemicals were of analytical grade and glass ware of borosilicate type.

Methods:

The determination of total phenolic content, total flavonoid content, total antioxidant and ferric chloride antioxidant reducing activity was carried out by spectrophotometry using Shimadzu UV-vis spectrophotometer.

Preliminary Phytochemical analysis^20-23:

The unani herbal formulations Habb-e-Suranjan and Habb-e-Azaraqi were subjected to qualitative chemical screening for identification of various classes of active chemical constituents by following standard methods.

Determination of total phenolic content:

The amount of total phenolics in extracts was determined according to the folin-ciocalteu procedure^24-25

Total flavonoid assay:

Total flavonoid content was measured by the aluminium chloride colorimetric assay^26-28

Determination of total antioxidant capacity:

The total antioxidant capacity of the food stuffs was evaluated by phosphomolybdenum method^29-30

Determination of ferric chloride reducing anti-oxidant capacity:

The total antioxidant activity can be measured by the ferric reducing antioxidant power assay (FRAP)^31-35

RESULTS:

Physical characters of formulations:

Table 1: Type of dosage form, pack size and Nature of herbal formulations Habb-e-Suranjan and Habb-e-Azaraqi

Name of the product	Type of Dosage form	Pack size	Nature
Habb-e-Suranjan	Solid Dosage form	50mg tablet	greenish brown in colour, characterstic agreeable odour
Habb-e-Azaraqi	Solid dosage form	50mg tablet	Reddish brown in colour, characterstic agreeable odour

Preliminary phytochemical screening of Unani formulations Habb-e-Suranjan and Habb-e-Azaraqi

Table 2: Phytochemical Screening of Habb-e-Suranjan and Habb-e-Azaraqi

CHEMICAL TESTS	Habb-e-Azaraqi	Habb-e-Suranjan
Test for tannins
Ferric chloride test	+++	+++
Lead acetate Test	+++	+++
Gelatin Test	+++	+++
Bromine water test	+++	+++
Acetic acid solution test	---	---
Potassium dichromate test	---	---
Dil. Iodine solution test	---	---
Dil. HNO₃ test	---	---
Dil. NH₄OH & Potassium ferricyanide solution test	---	---
Dil. KMNO₄ test	---	+++
Test for Saponins
Frothing test	---	+++
Test for Alkaloids
Dragendorff’s reagent	+++	+++
Mayer’s reagent	+++	+++
Hager’s reagent	+++	+++
Wagner’s reagent	+++	+++
Tannic acid Test	+++	+++
Tests for carbohydrates
Molisch’s test	+++	+++
Barfoed’s test	+++	+++
Standard test for combined reducing sugars	+++	+++
Standard test for free reducing Sugar (Fehling’stest).	+++	+++
Test for ketones	+++	+++
Test for pentoses	+++	+++
Test for phlobatannins (HCl test)	---	---
Test for cardiac glycosides
Baljet Test	---	---
Raymonds Test	---	---
Test for steroids
Liebermann-Burchard’s test	+++	+++
Salkowski’s test	+++	+++
Test for flavonoids
Shinoda’s reaction test	+++	+++
Pew’s test	+++	+++
Lead acetate test	+++	+++
Addition of sodium hydroxide	+++	+++
Test for anthraquinones
Borntrager’s Test	---	---

+++ Positive --- Negative

Determination of Total phenolic content

Figure1: Calibration curve of gallic acid

Table 3: Total phenolic content of Unani formulations

Sr. No.	Test Sample	Absorbance	Mean±SEM	Gallic acid equivalents
1	Habb -e- Suranjan	0.396	0.398±0.0015	70.34µg/ml
		0.401
		0.399
2	Habb -e- Azaraqi	0.231	0.253±0.0118	53.66 µg/ml
		0.272
		0.256

Values are expressed in mean ± SEM of 3 replicates

Table 4: Total Flavonoid Contents of Unani Formulations

Sr. No	Unani formulation	Absorbance	Mean ± SEM	Total flavonoidal content µg/ml
1.	Habb -e- Suranjan	0.135	0.148 ± 0.0701	103.49 µg/ml
		0.146
		0.163
2.	Habb -e-Azaraqi	0.103	0.112 ± 0.0530	78.32 µg/ml
		0.121
		0.112

Values are expressed in mean ± SE of 3 replicates

Determination of Total flavonoid content

Figure 2: Calibration curve of Quercetin

Figure 3: Calibration Curve of Ascorbic Acid

Determination of Total antioxidant capacity by phosphomolybdenum method

Table 5 : Total Antioxidant Capacity of Unani Formulations by phosphomolybdenum method

Sr. No.	Test Sample	Absorbance	Mean ± SEM	Ascorbic acid equivalent (μg/ml)
1.	Habb -e- Suranjan	0.575	0.575±0.2799	524.13 μg/ml
		0.454
		0.696
2.	Habb-e- Azaraqi	0.772	0.739±0.3532	679.54 μg/ml
		0.624
		0.821

Values are expressed in mean ± SE of 3 replicates

Determination of Ferric Reducing Anti-Oxidant Power (FRAP) Assay:

Table 6 : Total Antioxidant Capacity of Unani Formulations

Sr. No.	Test Sample	Absorbance	Mean ± SEM
1.	Habb -e- Suranjan	0.185	0.184±0.2799
		0.183
		0.184
2.	Habb-e-Azaraqi	0.375	0.376±0.3532
		0.379
		0.373
3.	Standard (Vitamin-C)	0.638	0.638±0.3532
		0.638
		0.638

DISCUSSION:

The potent antioxidant property of plants and their formulations has been centre of attraction for researchers throughout the world to prevent or reduce the progression of chronic disease and also due to economic viability and lack of side effects. One among many systems of traditional herbal systems unani medicines are formulated generally from natural resources and are well tolerated and this system of medicine needs to be scientifically explored for the benefits of mankind.

In the present study, the two unani formulations Habb -e- Azaraqi and Habb -e- Suranjan anjan were screened for phytochemicals, quantified for Phenolic compounds, flavonoids and assayed for total antioxidant capacity, ferric reducing antioxidant power to prove one of the mechanism of their therapeutic action is through free radical scavenging activity. These formulations have been using since many centuries in unani system of medicine for treating rheumatism, arthritis, gout, sciatica, back ache, facial paralysis, muscle pain and lumbago. The above-mentioned conditions involve excessive free radical generation leading to tissue damage, inflammation, pain and subsequent complications.

Phytochemical screening of above formulations (Table 1) revealed presence of phenolic compounds, flavonoids, alkaloids, tannins, phytosterols, Phenolic compounds and carbohydrates. A quantitative study of Phenolic compounds and flavonoids were performed by folin ciocalteau (FC) colorimetery and Aluminum chloride method respectively and Table 3 & 4 shows the total phenol and flavonoid contents of the formulation that were expressed in terms of gallic acid equivalent (GAE) and quercetin equivalent.

The present study also involved determination of total antioxidant capacity and ferric reducing antioxidant power for the unani formulations using phosphomolybdate method and metal chelating method. The antioxidant capacity of Habb -e- Azaraqi and Habb -e- Suranjan were found to be 679 and 524 ascorbic acid equivalents Table 5 and ferric reducing antioxidant power assay Table 6 exhibited IC₅₀ value. The formulations Habb -e- Azaraqi and Habb -e- Suranjan were found to be good activity.

ACKNOWLEDGEMENTS:

The authors are thankful to Prof. Dr. M. Srinivasa Murthy, Principal, Vignan Institute of Pharmaceutical Sciences, Deshmukhi, Hyderabad for providing facilities to carry out the work

FUNDING:

This research did not receive any specific grant from funding agencies in the public, commercial, or not for profit sectors.

CONFLICTS OF INTEREST:

The authors declare no conflicts of interest.

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Received on 19.01.2021 Modified on 11.08.2021

Research J. Pharm. and Tech 2022; 15(9):4111-4114.

DOI: 10.52711/0974-360X.2022.00690

0.135

0.146

0.163

0.103

0.121

0.112