INTRODUCTION:

Indigenous herbs in the form of oils from ancient period are vitally used in natural medications to treat various illness. These oils are administered either individually or combined with other powders, pills, etc.¹ In early days of civilization, people trusted herbal oil formulation as one of the best methods of application for therapeutic purposes and found effective remedies.^2,3 Massage therapy and Aroma therapy using herbal oils are still used in Siddha and Ayurvedic medicines in India.^4,5

Psidium guajava Linn, belongs to family Myrtaceae have been analysed for its pharmacological activity namely anti-microbial⁶, antioxidant⁷, anti-plasmodial⁸, antidiabetic⁹ etc. Psidium guajava is used in treating many diseases like rheumatic pain, wounds, reducing fever, hypertension and ulcers.¹⁰ The therapeutic property of the guava leaf extract are due to the bioactive compounds namely polyphenols, flavonoids, and terpenoids.¹¹ This article focusses on formulating the guava leaf oil and study its general, physical and chemical characteristics for future applications in treatment.

MATERIALS AND METHODS:

Preparation of Herbal oil:

The leaves were collected, washed with distilled water, and then left to dry. After being dried and ground into a powder, the Pg leaves were mixed with an adequate quantity of coconut oil (10g/100ml) and then preserved in glass bottles. The bottles were covered with muslin linen and left in the sun for 30 days so that the oil could absorb the herb extract. The treated samples were subjected to various characterisations.

General Evaluation:

The color and odor of the Guava leaf oil was observed.

Physical Evaluation:

Primary skin irritation test:

A small amount of Pg oil was applied to the test location, and observed for primary skin irritation on the forearm for approximately 1 centimetre. For 3-4hours, the test site was watched for erythema and edoema.¹²

Acid value:

Sample preparation involved taking 10 millilitres of oil, mixing it with 25 millilitres of ethanol-ether mixture (1:1ratio), and shaking the mixture. Using phenolphthalein as an indicator, 0.1 molar KOH was collected in a burette and titrated against the processed oil sample. It was computed how much KOH the oil used.

Saponification value:

One millilitre of carefully weighed oil was added to a 250mL conical flask, and ten millilitres of a 2:1 ethanol:ether mixture were added after that. The mixture was mixed with 25 millilitres of 0.5N alcoholic KOH, and the flasks were heated for 30 minutes before cooling. Using phenolphthalein indicator, the solution obtained after cooling was titrated against 0.5N HCl. The amount of KOH consumed was calculated in milligrammes.

Ester value:

The ester value of an oil is determined by taking the milligrammes of potassium hydroxide required to saponify the esters in one gramme of the oil. the ester value of an oil based on its acid value and saponification value (Ester value = Saponification value - Acid value)

pH:

The pH of herbal hair oil was determined using pH meter.

Specific gravity:

The specific gravity of the herbal oils was measured using specific gravity strips.

Chemical Characterization:

By pelleting the guava leaf oil with KBr (Nicolet 400), Fourier transform infrared (FT-IR) spectra were obtained, with peaks found in the 400–4000 cm−1 range. Additionally, the guava leaf oil was submitted to a mass spectrum analysis using the National Institute of Standards and Technology (NIST) database and a qualitative analysis utilising the JEOL GCMATE II GCMS to identify phytocompounds.

Antioxidant Activity- DPPH radical scavenging activity:

The method outlined in Braca et al. ¹³ was used to test the DPPH free radical scavenging activity. Ascorbic acid was used as a positive control standard ¹⁴.

Anti-inflammatory Activity - In vitro anti-arthritic activity:

In vitro anti-arthritic effectiveness was demonstrated using the protein denaturation method. ^15–17 The turbidity of the reaction mixture was measured at 600 nm. Protein denaturation inhibition percentage was measured based on the turbidity and Diclofenac sodium (250μg/ml) was taken as a positive control

RESULTS AND DISCUSSION:

Physical Evaluation:

The guava leaf after subjecting to physical evaluation showed the characteristic features in table 1.

Pg oil was able to effectively scavenge the free radicals in different concentrations in a dose dependant manner in all the assays and this action of antioxidants on DPPH radical scavenging was thought to be due to their hydrogen donating ability.

Table 2 GCMS peak table of Psidium guajava leaf oil

Peak	R.Time	Area	Area %	Name
1	3.042	3335643	0.67	BENZENE, ETHYL-
2	3.563	4843952	0.97	2-PENTANETHIOL, 2-METHYL
3	3.888	2742266	0.55	3-NONANONE
4	4.799	5118032	1.02	3-HEXEN-2-ONE
5	15.361	5297317	1.06	2-DECENAL, (E)
6	35.855	4050447	0.81	HEXADECANOIC ACID, 2-HYDROXY-1,
7	36.038	3534896	0.71	HEXADECANOIC ACID, 2-HYDROXY-1,
8	39.618	8624218	1.73	Octadecanoic acid, 2-hydroxy-1,3-propanedi
9	40.489	1688073	0.34	Oleic Acid
10	42.601	4885515	0.98	Dodecanoic acid, I-(hydroxymethyl)-1,2-eth
11	42.757	10507784	2.10	HEXADECANOIC ACID, 2-HYDROXY-1
12	45.159	8644382	1.73	ETHYL (9Z, 12Z)-9,12-OCTADECADIENO
13	45.238	23421940	4.68	9-Octadecenoic acid, 1,2,3-propanetriyl ester
14	45.315	2603554	0.52	12-TRICOSANONE
15	45.582	5543469	1.11	Glycidol sterarate
16	46.571	1595487	0.32	Dodecanoic acid, 1-(hydroxymethyl)-1,2-eth
17	46.848	1518112	0.30	Tetracosamethyl-cyclododecasoiloxane
18	48.580	201348008	40.27	2-LAURO-1,3-DIDECOIN
19	48.725	195512948	39.11	2-LAURO-1,3-DIDECOIN
20	48.885	5135245	1.03	GLYCEROL TRICAPRYLATE
		499951288	100

The GCMS analysis revealed that the guava leaf oil contains phenols, Dodecanoic acid responsible for anti-histaminic and anti-arthritic activity of the oil. 40.27% peak area was attributed by the compound 2-lauro-1,3 – Didecoin which exhibits several biological activity of the Pg oil like anti-dandruff, anti-microbial, antioxidant and anti-allergic activities²². These bioactive components supports the guava leaf oil in treating various ailments.

Antioxidant Activity- DPPH Radical Scavenging Activity:

The DPPH radical scavenging activity of Psidium guajava leaf oil was analyzed and its percentage inhibition was listed in the table 3. At varying doses, Psidium guajava oil demonstrated efficient scavenging of the free radicals in a dose-dependent manner. The DPPH free radical scavenging activity is a widely recognized model for mitigating lipid oxidation^23,24.

Anti-inflammatory Activity - Protein Denaturation Study:

According to the table, Pg oil in this investigation inhibited the denaturation of bovine serum albumin in a dose-dependent way.²⁵

Table 3: DPPH Radical Scavenging Activity by Psidium guajava leaf oil

Conc (µg/ml)	Ascorbic acid % Inhibition	*Psidium guajava* % Inhibition
50	81.2 ±2.38	70.61±1.24
100	89.56±2.21	75.43±1.21
200	91.93±0.35	81.24±1.35
300	95.49±1.65	83.11±1.36
400	101.53±2.69	85.26±1.45
500	102.77±2.41	91.43±1.58
IC ₅₀(µg/ml)	143	266

Table 4: Anti-inflammatory activity of Psidium guajava leaf oil

S. No.	Conc. (µg/ml)	Pg oil	Diclofenac Sodium
1	50	25.92
2	100	37.62
3	200	49.93	87.69
4	300	57.62
5	400	62.71
6	500	70.37
7	1000	77.12

CONCLUSION:

Currently researchers started exploring the traditional herbs and formulating them in different forms for its therapeutic purposes. One among such indigenous herb is Psidium guajava Linn (Guava). This study brings out the general, physical and chemical characteristics of Guava leaf oil and found that it is a potent herbal oil with antioxidant and anti-inflammatory property and can be used in treating various ailments. The oil can be further studied invitro and invivo and can be applied in various medical applications.

ACKNOWLEDGEMENT:

I express my sincere thanks to the management of Sathyabama Dental College and Hospital and SRM Dental College and Hospital, Ramapuram campus for providing great support in completing this research work.

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Received on 18.06.2024 Revised on 12.02.2025

Accepted on 16.07.2025 Published on 01.12.2025

Available online from December 06, 2025

Research J. Pharmacy and Technology. 2025;18(12):5651-5654.

DOI: 10.52711/0974-360X.2025.00816

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