Herbal medicines had glorious past and promising future throughout the world. Hence it is necessary to identify and characterize the crude drugs well before the use. This can be easily and reliably done by the Pharmacognostic study. Pharmacognosy is the simultaneous application of various scientific disciplines with the object of acquiring knowledge of drugs from every point of view. Ixora coccinea Linn. is a shrub commonly known as Raktaka/ Rukmini/ Tetti etc. It belongs to family Rubiaceae.1 It is much branched shrubs, branch lets somewhat compressed. Leaves are sessile, subcordate and amplexicaul at base, elliptic or oblong, obtuse, apiculate. Flowers numerous in dense, corymbiform, sessile or pedunculated cymes.2 Ixora coccinea is native to India. The root, stem, leaves and flowers are used to treat various ailments in Indian traditional system of medicine. The use of flower in medicine is less known although it is mentioned in Ayurveda. The flowers of raktaka are used in dysentery, dysmenorrhoea, leucorrhoea, haemoptysis, catarrhal bronchitis, ophthalmopathy, sores, hypertension, anaemia and ulcers.3-4 It is also used as a remedy for skin diseases, colic flatulence, wounds, indigestion and as antiseptic.5 The chemical composition of Ixora coccinea flower were intensively investigated in previous studies. The anthocyanins were extracted and studied in detail by HPTLC.6 Ixora flower has been confirmed as a natural pH indicator in all acid base titrations.7 The new triterpenoid, Ixoroid was isolated from the Ixora flower with spectral evidence.8 The pharmacological activities of flower was reported to have antioxidant, antitumor, wound healing, hepatoprotective and as chemoprotective.9-11 However, the information regarding identification of the flower parts are not been carried out. As this flower has huge therapeutic potentials it needs to be explored and brought into world of herbal medicine hence, the current study was done. A detailed investigation on macroscopic, microscopic characters, histochemistry, physicochemical parameters, powder study, fluorescence analysis and preliminary phytochemical screening of the flower parts were done. These will help in identification and standardization of said drug.
MATERIAL AND METHODS
Collection of plant material
Authentic flower sample were collected from wilderness areas of Gorai and Dapoli, Maharashtra, India. The flower was collected in the month of January early morning. The sample was authenticated for its botanical identity from the Botany department, Mithibai College, Vile Parle (W), Mumbai, India. After collection some of the flowers were preserved in Formalin: Acetic acid: Alcohol solution. Remaining flowers were dried after removing stamens, style and made into powder. It was sieved through British Standard Sieve, meshes number 85 and stored in air tight bottles.
Pharmacognosy of the entire flower was carried out using standard methodology.
The entire flowers were studied for its morphological characters using appropriate techniques.12
Transverse hand cut sections of corolla, corolla tube and calyx with ovary were taken and made permanent with suitable stains.13 Quantification and photomicrographs were taken of the permanent preparations. The cell contents were measured using stage and ocular micrometer.14
The histochemical studies for the cell content were done by staining the hand cut sections of corolla, corolla tube, and calyx passing through ovary with different reagents.15
(d) Powder study
The dried powder of flower was treated with chloral hydrate solution followed by staining in 1% safranin for 5-10 mins and mounted in 50% glycerine.16
(e) Proximate analysis
The physicochemical parameters like ash values (total ash, water soluble ash and acid insoluble ash) and extractive values (water and alcohol extractive values) were established using powdered drug.17
(f) Fluorescence analysis
The fresh flower of Ixora coccinea is sessile, actinomorphic, bisexual, hypogynous and tetramerous. The calyx is with 4 sepals, short, triangular, persistent, gamosepalous measuring 0.3 - 0.35 – 0.4 cm in length. Corolla has four petals, oblong to ovate, gamopetalous, 0.5 – 0.7 cm in width; corolla tube is long scarlet orange colour, 3 – 3.2 – 3.5 cm in length. Androecium consists of 4 stamens, alternipetalous, extrose, anthers basifixed. The gynoecium is bicarpellary, syncarpous, bilocular ovary with basal placentation, bifid stigma. (Figure 1-2)
Organoleptic study of fresh flower: Colour – Scarlet orange, odour – characteristic, Taste – characteristic.
T.S of flower passing through corolla tube
Outer epidermis of corolla tube is single layered, compactly arranged, tangentially elongated cells measuring 90 - 105µm in length. It is covered with thick waxy cuticle and interrupted by unicellular trichomes. The cells below the lower epidermis are larger and filled with anthocyanin pigments. Mesophyll region consists of 8-9 layers of polygonal parenchymatous cells measuring 45 – 60 - 75µm in diameter. This region is embedded with vascular bundles consisting xylem on the inner side and phloem on the outer side. The cells are filled with abundant oil globules, starch grains and few anthocyanin filled cells. Inner epidermis is similar to that of outer epidermis. The epidermis is interrupted with stomata and very few smaller trichomes. (Figure 3)
T.S of flower passing through corolla
Outer epidermis of corolla is single layered tangentially arranged 89 - 108 µm in length. It is covered with thick cuticle and few unicellular trichomes. The cell below the outer epidermis is larger and is filled with anthocyanin pigments. Mesophyll region consists of 8-9 layers of closely packed parenchymatous cells 75 - 90 – 120 µm in diameter, with vascular bundles embedded at intervals. The parenchymatous cells are filled with oil globules and starch grains. Inner epidermis is similar to outer epidermis; it also shows a layer of anthocyanin filled cells above the inner epidermis. Trichomes are very few in number. (Figure 4)
T.S of flower passing through calyx and ovary
T.S. of Calyx: The outer epidermis of calyx is single layered, compactly arranged, tangentially elongated cells 30 - 45 - 70µm in length and covered with cuticle. It is interrupted with numerous unicellular trichomes and stomata. The outer epidermis of calyx is followed by a single layer of anthocyanin filled cells. Mesophyll region consists of 9 -10 layers of polygonal parenchymatous cells 105 – 120 – 135 µm in diameter. The mesophyll region is interrupted with vascular bundles. The cells are filled with oil globules, starch grains and calcium oxalate crystals. The inner epidermis of calyx is single layered with the absence of trichome and stomata.
T.S. of Ovary: The calyx surrounds the ovary. The ovary is bicarpellary. Both the ovaries are surrounded by an intact epidermis which is single layered and covered with cuticle. The central region of ovary is made up of parenchymatous cells closely arranged. The two ovaries are connected by a connective tube made of parenchymatous cells. (Figure 5)
The Ixora coccinea flower powder
The dried flower powder is scarlet orange in colour. The flower powder on treatment with chloral hydrate solution followed by staining in 1% Safranin for 5-10 mins and mounted in 50% glycerine exhibited fragments of parenchymatous cells, anthocyanin filled cells, paracytic stomata, annular vessels, starch grains, calcium oxalate crystals and oil globules. (Figure 6)
The histochemical analysis using various reagents showed the presence of primary and secondary metabolites like starch, lipids, proteins, tannins, saponins, glycosides and oil globules respectively.
The physicochemical constants of flower revealed total ash content 3.1%, water soluble ash content 1.25% and acid insoluble ash content 0.2%. Thus, the acid insoluble ash value states the presence of least amount of silica in flower powder. The extractive value of water is 12% and ethanol 11.25% the above extractive values determine that more chemical constituents are soluble in the water.
The fluorescence analysis of Ixora coccinea flower powder with different chemicals exhibited various colours in the UV light. The results are depicted in Table 1.
The preliminary qualitative phytochemical screening of alcohol and water extract of Ixora coccinea dry flower revealed the presence of terpenoids, flavonoids, anthraquinone glycoside, cardiac glycoside and tannins. The saponin was reported only in water extract. Table 2.
DISCUSSION AND CONCLUSION
Pharmacognosy is the first step in deciding the status of a plant organ as a crude medicine before including it in herbal pharmacopoeia. It is important to establish pharmacognostic parameters for the detection of adulterant or substitute. The current research work focuses on, the pharmacognostic investigations of Ixora coccinea L. flower. The said flower is used by traditional and local people without standardization and has obtained good results. Macroscopic observations of fresh flower such as persistent calyx, scarlet orange corolla with corolla tube, the stamens alternipetalous and gynoecium bicarpellary with basal placentation are useful for gross identification of the flower. Anatomical features like unicellular trichomes, parenchymatous cells filled with anthocyanin pigments, starch grains, calcium oxalate crystals and oil globules are of significance in recognition of floral parts. The corolla tube differs from the corolla in presence of anthocyanin filled cells below outer epidermis and above inner epidermis as well as in the mesophyll region. The calyx shows the presence of paracytic stomata on the outer epidermis and also the presence of calcium oxalate crystals. The paracytic stomata are restricted to inner epidermis of corolla and corolla tube. These microscopic characters can be used as a parameters in standardization of Ixora flower. Authentication of powdered drug can be reliably done on the basis of diagnostic characters of paracytic stomata, oil globules, starch grains, anthocyanin filled cells, calcium oxalate crystals and annular vessels. Fluorescence analysis will be useful in identification of Ixora coccinea flower in powder form. All physicochemical parameters like ash and extractive values are also of help in detection of adulterants. The qualitative phytochemical and histochemical screening revealed the presence of diverse types of phytochemicals namely, alkaloids, tannins, terpenoids, flavonoids, saponins, essential oil etc. They give clue about therapeutic potential of the drug. In brief, all these findings are highly essential for the drug manufacturers in thorough assessment of quality drug and its efficacy.