INTRODUCTION:

Medicinal plants accumulate variety of active principles or secondary metabolites which are used to treat various human or animal diseases¹. The genus Hypericum consists of about 400 species, of which ten morphologically and chemically distinct species are grown in Central Europe². Hypericum oblongifolium, belonging to the family Hypericaceae, is 60-120cm in height, an erect evergreen shrub, that grows on Khasia Hills at an altitude of 5000–6000 ft, and common in China, and in the Himalayan region³. This plant has been traditionally used for the treatment of various conditions as sedatives, to treat external wounds, gastric ulcer, remedy for sting of bees and dog bites, hepatitis, nasal hemorrhage, antiseptic and as antispasmodic⁴. H. oblongifolium has also been reported for lowering of blood pressure, bronchodilator, anti-glycation, antispasmodic, anti-ulcer, chymotrypsin and urease inhibition, antioxidant, anti-lipid peroxide inhibition, antidepressant, anti-proliferative activities, wound healing, anxiolytic, antiviral and antimicrobial activities³. The phytochemical analysis showed that H. oblongifolium contains flavonoids, saponins and tannins⁵, whereas the other chemical compounds reported in H. oblongifolium species include flavonoids like quercetin, rhamnetin, myricetin, kaempferol, triterpenes like hyperinols A and B, xanthones like hypericorin-C, hypericorin-D, 3,4-dihydroxy-5-methoxyxanthone and folicitin, a new compound with antioxidant activity⁴. For identification and collection of correct species there is need to establish the characteristic features of the plant.

MATERIALS AND METHODS:

Plant Material and Reagents:

H. oblongifolium leaves were collected from Kainchi, Nainital in August 2017. The specimen (voucher No. 126112) was identified by Dr. Kumar Ambrish (Scientist-D), at Botanical Survey of India, Dehradun and sample specimen was deposited in the herbarium of the BSI. The solvents used were ethanol, formic acid, methanol, gallic acid, and other lab reagents of analytical grade.

Macroscopy and Microscopy:

The leaves of H. oblongifolium (HOB) were separated from other parts, washed properly and dried under shade for further use. The macroscopic characters of the fresh leaves were noted: as color, odor, taste, shape, size, surface texture, arrangement and attachment of leaves⁶. The organoleptic characters of the leaves were observed, noted and photographs were taken. Microscopical structures of the leaf samples were observed by transverse section and powder microscopy^7,8. Small quantity of the leaf powder was placed on the surface of a glass slide, decolorized, stained and observed under the microscope to identify the characteristic structures^7,9.

Extraction:

Leaf sample was shade dried at room temperature and coarsely powdered before extraction. A known amount of plant sample was extracted by cold maceration method at room temperature using 70% ethanol (hydroalcoholic) as solvent. The extract was then concentrated by using a Rotary vacuum evaporator at 45 °C until a crude extract was obtained, after complete evaporation of the solvent extract was dried, and stored at 4-6°C till further use.

Determination of Physicochemical Parameters:

Physicochemical analysis includes determination of moisture content (loss on drying), determination of total ash, acid insoluble ash, water soluble ash, water soluble extractive and alcohol soluble extractive values. These parameters were evaluated as per the WHO guidelines for herbal plant materials^6,10,11.

Fluorescence analysis:

The shade dried leaf powder sample was treated with various reagents and emitted fluorescence was observed under visible and UV light (at 254nm). Reagents used for fluorescence analysis were dilute sodium hydroxide solution, iodine solution, aqueous ferric chloride solution, picric acid solution, acetic acid, ammoniacal solution, aqueous silver nitrate solution, concentrated sulphuric acid^12,13.

Phytochemical screening:

Crude ethanolic extract of H. oblongifolium was subjected to preliminary phytochemical screening to investigate the presence of phytochemicals like alkaloids, terpenoids, glycosides, steroids, tannins, flavonoids and saponins^6,14.

FTIR analysis:

Fourier Transform Infrared Spectrophotometer (FTIR) is the most powerful tool for identifying the functional groups present in the compounds¹⁵. The spectra were recorded in FTIR instrument of Perkin Elmer, with computer-based software. The chemical bonds in a compound can be determined by interpretation of the IR spectrum¹⁶. The FTIR analysis was performed for both powder sample and extract of H. oblongifolium leaf¹⁷. The IR analysis of the sample was performed over a wave number ranged from 4000 to 500cm^-1. The spectral data were compared with reference to identify the presence of functional groups in the sample¹⁸.

RESULTS AND DISCUSSION:

Macroscopy:

The leaves were observed as elliptical to oblong (so the name of species oblongifolium), 5-7.5cm in length and 2-3cm in width with acute apex and entire margin. The leaves were dark to light green in color with slight odor, taste slightly bitter. Leaves texture was smooth, and showed sessile or stalkless leaf attachment whereas the arrangement of the leaves was opposite-decussate, as shown in Fig.1.

Microscopy:

Transverse Section of the Leaves: The transverse section of H. oblongifolium leaf showed the presence of palisade layer below upper epidermis, collenchyma, sclerenchyma, and vascular bundles, their arrangements are shown in Fig.2.

Powder Characteristics of Leaf: The powder analysis of leaf showed the presence of beaded wall of epidermis, brownish epidermal cell, anomocytic stomata, elongated palisade cells, lignified xylem vessel (spiral), trichomes, and star shaped fibres as shown in Fig.3.

Physicochemical Characteristics:

The physicochemical characteristics such as loss on drying (0.34% w/w), water soluble extractive value (8.7 % w/w), and alcohol soluble extractive value (8.2% w/w) were measured. The ash values were found as 5.63 % w/w (total ash), 1.39% w/w (acid insoluble ash) and 4.241% w/w (water soluble ash). Total ash value of the sample is higher than other values because of the presence of oxalates, phosphates, carbonates, and silicates. Ash values indicate presence of inorganic components in the leaf sample, and called physiological ash. If ash values are more than the limit, it indicates presence of impurities which is called non-physiological ash. So, the determination of ash values and extractive values is important for the quality control of the genuine drug.

Fluorescence Analysis:

Fluorescence analysis of the powder material was performed under the visible and UV light (254nm), which further signifies its characteristic importance for identification. Results of fluorescence analysis of leaf powder using different reagents are shown in Table 1.

Phytochemical Screening:

The phytochemical screening of ethanolic extract showed presence of tannins, flavonoids, alkaloids, carbohydrates, glycosides and phenols; amino acid, protein and steroids were absent.

FTIR Analysis:

The FTIR spectra of leaf powder and leaf extract are shown in Fig 4 and 5 respectively. The peaks on the spectra and the probable functional groups present in the leaf powder of H. oblongifolium and leaf extract of the same are presented in Table 2 and 3 respectively. These spectra showed the presence of O-H, C=C and C-O bonds in both the leaf powder and the extract, so confirmed the presence of functional groups having such bonds like, hydroxyl group, acid group, alkene, ester or ether groups. Presence of theses groups in both the samples indicated that the functional groups remain intact during extraction process as these are responsible for the pharmacological activity of the plant.

Fig. 1: Macroscopic characters of H. oblongifolium leaves

Fig. 2: Transverse section (T.S.) of H. oblongifolium leaf

Fig. 3: Powder microscopy of H. oblongifolium leaves

Fig. 4: FTIR spectrum of H. oblongifolium leaf powder

Fig. 5: FTIR spectrum of H. oblongifolium leaf extract

Table 1: Fluorescence Analysis of H. oblongifolium Leaf Powder

S. No.	Sample treated with	Result
S. No.	Sample treated with	UV light (254 nm)	Visible light
1	Dilute NaOH	Brownish	Brownish
2	Iodine solution	Blackish	Yellowish
3	Ferric chloride	Brownish	Yellowish
4	Picric acid solution	Yellowish	Pale Yellow
5	Acetic acid	Brownish	Brownish
6	Ammonia solution	Yellowish	Yellowish
7	Silver nitrate solution	Blackish	Brownish
8	Concentrated sulphuric acid	Dark Brown	Brownish

Leaf powder of the drug was treated with different reagents on the glass slide and observed under UV and visible light

Table 2: FTIR Analysis of Powder Sample of H. oblongifolium Leaf

S. No.	Peak value (cm^-1)	Functional groups
1	3278.48	O-H; stretching
2	1605.97	C=C; stretching
3	1439.25	CH_3; bending
4	1017.52	C-O; stretching

wave no. (cm^-1)

Table 3: FTIR Analysis of H. oblogifolium leaf Extract

S. No.	Peak value (cm^-1)	Functional groups
1	3265.99	O-H; stretching
2	1599.1	C=C; stretching
3	1445.34	CH₃; bending
4	1060.3	C-O; stretching

wave no. (cm^-1)

CONCLUSION:

The data generated from the present study would help in the authentication of this plant both in fresh and dried powder form and to distinguish it from other species. The microscopic features and the standards observed would be useful for setting down pharmacopeial standards. Phytochemical screening, and FTIR analysis have shown the presence of different secondary metabolites which are mainly responsible for pharmacological activity of the plants. FTIR analysis is also useful in confirming the presence of functional groups in the extract by comparing with powder sample of the leaf. Finally we can conclude that these parameters will be helpful for identification and collection of the genuine species.

ACKNOWLEDGEMENTS:

The authors wish to thank Head, Department of Pharmaceutical Sciences, Faculty of Technology, Kumaun University Nainital, Uttarakhand, India for providing necessary facilities to complete this work.

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Received on 20.01.2025 Revised on 25.07.2025

Accepted on 19.11.2025 Published on 31.01.2026

Available online from February 07, 2026

Res. J. Pharmacognosy and Phytochem. 2026; 18(1):13-16.

DOI: 10.52711/0975-4385.2026.00003

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