Eugenol Content Research Articles

Quality control of marketed clove buds - reference to their quality and purity as per who guidelines

Two marketed clove buds were subjected to crude drug standardization by sampling from retail packages as per WHO recommended guidelines. The marketed clove products viz. brand-A and brand-B were standardized comparatively for various parameters like, powder fineness, foreign matter, ash values, volatile oil content, bitterness value, swelling index, foaming index, heavy metals, Mycological examinations, spectral analysis, qualification of Eugenol by TLC and quantification by HPLC. The results were compared with individual monograph limits specified in Indian Pharmacopoeia. The volatile oil content were estimated by Azeotropic distillation and were found to be 1.5ml and 1.2ml/10gms respectively for brand-A and brand-B against the limit 1.5 to 2.1ml. The UV spectra and ATR-IR spectra of both volatile oils were identical with the finger print of standard Eugenol oil. The foreign matter and ash value for brand-A and brand-B were 1.49%, 5.8% and 3.79%, 6%, respectively and brand-B doesn’t complies with the specified monograph limits (NMT 3%) for foreign matter. The Eugenol content in the volatile oil was quantitated by RP-HPLC method and was found to be 93.3.1% and 74.6% respectively for brand-A and brand-B against the standard monograph limit of 85-95%. It was noted that the brand-A was found to be superior quality and whilst brand-B disqualifies in standardization parameters such as Foreign matter, Volatile matter and Eugenol content. World Health Organization has emphasized the need to ensure quality control of medicinal plant products by applying suitable parameters and standards. In order to overcome certain inevitable shortcoming of the Pharmacopoeial monograph and other quality control measures, the present study explores the possible parameters of crude drug standardization for marketed clove buds with special reference to their quality and purity as per WHO guidelines.

The beneficial use of essential oils from buds and fruit of Syzygium aromaticum to combat pathogenic fungi of Panax notoginseng

To develop natural fungicides against Panax notoginseng disease, the antifungal effects of Syzygium aromaticum essential oils (EOs) against the pathogenic fungi causing P. notoginseng disease were investigated. The effects of EOs from S. aromaticum buds and fruits on the growth of pathogenic mycelia were determined with in vitro antifungal experiments using the Oxford cup method. The results suggested that S. aromaticum EOs had a strong inhibitory effect on the growth of pathogenic fungi. At a dose of 50 mg/mL, the inhibitory effects of S. aromaticum buds EOs was greater than those of S. aromaticum fruits. Then the chemical constituents of the S. aromaticum EOs were analyzed by GC–MS. The results indicated that the eugenol content was the highest, accounting for 57.95% of the total constituents in the S. aromaticum buds EOs and 39.26% in the S. aromaticum fruits EOs. The minimum inhibitory concentration (MIC) of EOs, the major constituents, and hymexazol were evaluated. In addition, the antifungal effect of their combination was also determined by the fractional inhibitory concentration (FIC). When the buds EOs were combined with hymexazol, they had a synergistic effect in inhibiting Pythium aphanidermatum, Botrytis cinerea, and Colletotrichum gloeosporioides. The combination of S. aromaticum fruit EOs with hymexazol showed an additive effect on Fusarium solani, P. aphanidermatum, and B. cinerea. The preventive effect of S. aromaticum bud EOs combined with hymexazol was up to 60.77% in in vivo experiments. Overall, S. aromaticum EOs had a broad spectrum of antifungal activities against pathogenic fungi influencing the growth of P. notoginseng, which would provide a scientific basis for the ecological control of P. notoginseng disease.

Effect of Zinc on the Growth and Essential Oil Composition of Ocimum gratissimum L.

Ocimum gratissimum, commonly known as Rama Tulsi is a rich source of aroma chemicals that find varied application in the food, flavor and pharmaceutical industry. A polyhouse study was conducted to determine the effect of zinc on the growth and essential oil composition of O. gratissimum L. The treatments of zinc were 600, 1000 and 1200 mg kg-1 in triplicate along with a control treatment. Zinc content was examined by AAS while hydrodistilled oil was analyzed by GC and GC-MS. The highest fresh weight and plant height was obtained from the plants grown in control soil. Ocimum gratissimum treated with Zn at a rate of 600 mg kg-1 accumulated greater concentrations of Zn. Thirty four, 36, 36 and 29 compounds constituting 93.62, 85.99, 93.60 and 9525 % of the total oil were identified in the essential oil of control, Zn600, Zn1000 and Zn1200 treatments respectively. The main components in the oil of plants grown in control soil were germacrene D (31.04 %), (E)-caryophyllene (9.00 %), (Z)-β-ocimene (8.67 %), α-copaene (8.48 %), eugenol (7.87 %) and caryophyllene oxide (7.03 %). The content of eugenol was found to decrease significantly from 7.87 to 1.99 % while that of maaliol which was absent in the control, increased to 17.65 % with 1200 mg kg-1 of Zn treatment.

Eugenol‐Derived Bismaleimide High Performance Resins and Composites Using Diisocyanate as Property Modifier

AbstractA new kind of high performance bismaleimide resin with good processability and improved toughness is synthesized by chemical modification of 4,4′‐bismaleimidodiphenylmethane (BMI) by eugenol (EG) and different contents of 4,4′‐diphenylmethane diisocyanate (MDI). MDI‐EG‐BMI resins exhibit good thermal stability for its 5% weight loss temperatures around 300 °C and its residue of 41.61% at 900 °C, which are much higher than those of EG‐BMI resin. Then, the carbon fiber‐reinforced MDI‐EG‐BMI composites are fabricated. The mechanical properties of the composites matrixed by MDI‐EG‐BMI resins are better than those by EG‐BMI resin. For carbon/MDI‐EG‐BMI composites, their glass transition temperatures are higher than 300 °C, and their flexural strength, moduli, and toughness are maintained at a range of 217.47–404.36 MPa, 35.12–48.49 GPa, and 1.16–2.63 MJ m−3 respectively; with the contents increasing of MDI in the resin formulation, the flexural properties first increase then decrease; comprehensively the composite with 30 wt% MDI has the best mechanical and thermal properties.

Pharmacognostic Evaluation and Pharmacological Validation of Sphaeranthus indicus DC

Sphaeranthus indicus is an important medicinal plant in Ayurveda which grows as weed in rice fields throughout India, Sri Lanka, Australia and Africa. The present study aimed for the pharmacognostical evaluation and pharmacological validation of this high-value medicinal species. All the pharmacognostical parameters were done as per the API guideline. A simple, rapid, sensitive and reproducible method was also developed for simultaneous HPTLC quantification of two bioactive compounds eugenol and β-sitosterol in S. indicus. The HPTLC was performed on silica gel 60 F254 by using toluene/ethyl acetate/glacial acetic acid (8:2:0.2) as a mobile phase for eugenol and β-sitosterol at Rf value 0.64 and 0.48, respectively. The content of eugenol (0.03%) and β-sitosterol (0.013%) was detected in dry weight per gram. In vitro activities viz. antioxidant, antidiabetic and anti-inflammatory were done to evaluate the pharmacological potential of S. indicus. Four antioxidant models viz. DPPH, ferric reducing power, 2-deoxyribose assay and antioxidant capacity were used to determine the free radical scanning. Antidiabetic and anti-inflammatory potential were determined by using starch–iodine colour assay and inhibition of protein denaturation model. IC50 value of S. indicus extract in DPPH and 2-deoxyribose assay was found (328.96 ± 0.003 µg/ml) and (30.36 ± 0.004 µg/ml), respectively. The present study will be helpful for quality check of the raw material and monitoring batch-to-batch consistency of herbal drugs, wherein S. indicus is used as an ingredient.

Effect of drying temperature and storage time on Ocimum gratissimum Linn. leaf essential oil from Central Highlands, Vietnam

Abstract Essential oil of Ocimum gratissimum Linn. leaf is widely used in folk medicine. Extraction yeild, compositions and antioxidant activity of clove basil essential oils could be affected by several factors including drying temperature and storage conditions. The present work evaluated the influence of storage conditions and drying temperature different of Ocimum gratissimum leaves on essential oils from Central Highlands, Vietnam. In this research, fresh leaves were preserved at 4°C for 10 days and dried and then stored at room temperature. Essential oil of Ocimum gratissimum L. was extracted by hydrodistillation and steam distillation methods. The composition of essential oil was determined by using GC-MS, antioxidation activities were conducted by the DPPH method. Drying the material at 45°C only 1% Decrease extraction yield, the extraction yield of essential oil at 45°C is still high (2.5%); the color of the oil when the material is fresh in yellow, characteristic odor, drying at 45°C is pale yellow, mild characteristic odor; Antioxidation resistance change is negligible; Drying the material at 45°C, the content of eugenol (77.389%) increased 3 fold compared with fresh material (24.84%), while the drying loss of some components small proportion in oil.

Effects of the Syzygium aromaticum L. extract on antioxidation and inhibition of matrix metalloproteinase in human dermal fibroblast

Objective: To investigate cosmetic potential of Syzygium aromaticum L. (S. aromaticum L.) and to determine its antioxidant and anti-wrinkling effects. Methods: Using high-performance liquid chromatography, eugenol component was quantitated. The antioxidant activity of S. aromaticum L. was analyzed by 2,2-diphenyl-1-picrylhydrazyl radical scavenging and superoxide dismutase like activities. To determine cell viability, elastase and matrix metalloproteinase-1 (MMP-1) activity, human dermal fibroblasts (HS68) were treated with S. aromaticum L. The inhibitory effect of S. aromaticum L. on tumor necrosis factor alpha induced MMPs expression in HS68 was analyzed by realtime-PCR. Results: The eugenol content was confirmed in S. aromaticum L. S. aromaticum L. was observed to have high 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and superoxide dismutase like activity. S. aromaticum L. had no cytotoxicity against the HS68 and dose-dependently increased elastase inhibition. Moreover, S. aromaticum L. significantly decreased MMP-1 content and inhibited gene levels of MMP-1, MMP-2, MMP-3 and MMP-9. Conclusions: The findings suggest that S. aromaticum L. has great potential as a cosmeceutical ingredient with antioxidant and anti-wrinkling effects.

<i> Syzygium aromaticum</i> L. (Clove) Essential Oil as a Reducing Agent for the Green Synthesis of Silver Nanoparticles

The present investigation details a green synthesis of silver nanoparticles (AgNP) using the essential oil of Syzygium aromaticum L. (clove) as reducing agent, which is a matrix with a high content of eugenol, an important compound for the reducing action of silver nitrate. The synthesis of AgNP was performed at different pH conditions (pH 7, 8, 9 and 10), and was monitored by UV-Vis Spectroscopy, Dynamic Light Scattering (DLS) and Transmission Electron Microscopy. The synthesized nanoparticles presented characteristic Surface Plasmon Resonance bands with maximum absorbance between 405 and 460 nm. The DLS analysis revealed particle sizes from 31 to 72 nm and zeta potential between -30.1 and -50.8 mV indicating good stability against the agglomeration of the particles in solution. The micrographs obtained by TEM showed different particle shapes and a predominance of spherical-shaped nanoparticles, and average size ranging from 27 to 94 nm. The clove-based silver nanoparticles were efficient in controlling the growth of Escherichia coli and Staphylococcus aureus bacteria, and the minimum inhibitory concentration ranged from 60 to 100 μL/mL. This study highlights the feasibility of clove essential oil as an alternative for the synthesis of silver nanoparticles by a simple, inexpensive and eco-friendly method.

Evaluation of the effects of temperature and pressure on the extraction of eugenol from clove (Syzygium aromaticum) leaves using supercritical CO2

In this study, the effect of operational conditions on the extraction of clove leaves using CO2 on eugenol yield and content was investigated. The conditions used in the supercritical extraction with CO2 included temperatures of 40, 50, and 60 °C and pressures of 150, 185 and 220 bar. The highest yield (1.08 wt%) and highest eugenol content were obtained at 40 °C and 220 bar. The main components identified in the clove extract were eugenol (29.73%), chavicol (13.31%), n-pentacosane (11.66%), hexacosanal (11.37%), and vitamin E (9.30%). The extracts were evaluated for their antioxidant capacities by the DPPH and FRAP methods, and quantification of the phenolic compounds was done by the Folin-Ciocalteu method. The best results among the conditions were obtained at the lowest pressure and temperature, 150 bar and 40 °C. The Sovová model satisfactorily described the extraction kinetics in all the studied conditions.

Chemical Composition of Leaf Essential Oils of Four Cinnamomum Species and Their Larvicidal Activity Against Anophelus sinensis (Diptera: Culicidae)

Essential oils from Cinnamomum camphora, Cinnamomum japonicum, Cinnamomum szechuanense and Cinnamomum subavenium were investigated for their chemical compositions and larvicidal activities against Anopheles sinensis larvae. The results showed that a total of 51 chemical compounds were identified from the four essential oils. The major ingredients of C. subavenium essential oil were engenol (67.73%) and engenol acetate (13.19%), while C. szechuanense, C. camphora and C. japonicum was 1,8-cineole, accounting for 42.90%, 40.91% and 24.62% in total oil, respectively. More importantly, engenol and engenol acetate were only detected from C. subavenium essential oils, suggesting that the chemical composition of C. subavenium essential oil was quite different from those of C. camphora, C. szechuanense and C. japonicum. Cluster analysis also confirmed the significant difference between C. subavenium and the other three essential oils. The four essential oils all acted against A. sinensis larvae in a dose-dependent manner. Among them, C. subavenium essential oils have the strongest larvicidal activity with the lethal concentration 50 (LC50) of 79.63 μg/mL, while the LC50 of other essential oils were 96.81 μg/mL (C. szechuanense), 109.40 μg/mL (C. camphora) and 117.80 μg/mL (C. japonicum), respectively. Comparative analysis of the chemical compositions and the larvicidal activities of the essential oils showed that the higher larvicidal activity of C. subavenium essential oil was probably due to its high eugenol and engenol acetate content. These oils, especially C. subavenium essential oil, should be promising alternatives to existing larvicides, or could be integrated into larval source management compounds for A. sinensis control.

Variation in Growth and Development, and Essential Oil Yield between Two Ocimum Species (O. tenuiflorum and O. gratissimum) Grown in Georgia

The use of medicinal plants in the United States is increasing. Holy basil (Ocimum tenuiflorum L. and Ocimum gratissimum L.), a medicinal herb native to India, has become increasingly popular for its therapeutic benefits. Traditionally, holy basil has been used to promote longevity by reducing stress and restoring balance to the body. Because it is easy to grow and adapts to a wide range of growing conditions, there is great potential for holy basil production in the southeastern United States. The purpose of this study was to evaluate holy basil varieties for harvestable weight and essential oil content. Fourteen varieties of holy basil were grown during the 2015 and 2016 seasons and compared. The main active compound in the holy basil essential oil fraction, eugenol, was quantitated and compared for each variety, because it is believed to be responsible for many of the health-promoting effects. Overall, there were significant differences in harvestable weights and essential oil yields among the varieties, and a significant effect of growing season. The eugenol content was highly variable among the varieties examined, with higher eugenol contents in 2016 than in 2015. The variety that had the overall highest yield, essential oil content, and eugenol concentration was PI 288779, a USDA accession, suggesting its use in future breeding research.

Pertumbuhan dan Kandungan Minyak Atsiri Tanaman Selasih (Ocimum basilicum L.) pada Naungan dan Dosis Pupuk Fosfat yang Berbeda

ABSTRACT<br /><br />Basil (Ocimum basilicum L.) is a plant that has great potency to be developed. Almost all parts of the plant can be utilized. This study aimed to determine the effect of shade and phosphate fertilizer on growth and essential oil content in basil. This research was conducted at the experimental field of Agrotechnology Departement, Faculty of Agriculture, University of Trunojoyo Madura, during January-April 2015. The research was arranged in a split plot design with 3 replications. Main plot was shading i.e., without shading and 25% shading. The subplots were phosphate fertilizer rate, i.e., 0, 1.5, 3.0, and 4.5 g SP36 per plant that was planted in polybags. The results showed that plants treated with 25% of shading and applied with phosphate 3 g per plant had the highest total dry weight, production of essential oil and eugenol. Shading of 25% give the best results on plant height, leaf area and total fresh weight of the plant. Application of phosphate fertilizer 3 g per plant give the highest content of chlorophyll (chlorophyll b and total chlorophyll), while the highest essential oil and eugenol content of leaves is obtained from application of phosphate fertilizer 3 and 4.5 g per plant. Shading and phosphate fertilizer application did not affect leaf number. <br /><br />Keywords: chlorophyll, eugenol, leaf, phenylpropanoids, production<br /><br />

Study on the bacterial biofilm of Streptococcus pneumoniae by clove oil

Objective To study the effect of clove oil on bacterial biofilm of Streptococcus pneumoniae. Methods The components of clove oil were determined by GC-MS mass spectrometry. The suspension of streptococcus mutans was prepared to carry out the germicidal test, and to determine the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of clove oil. The effects of different concentrations of clove oil on the formation of biofilm were observed by laser confocal microscopy, and the average fluorescence intensity of live bacteria and dead bacteria in biofilm was recorded. Results Determination of clove oil 8 components were analyzed by GC-MS mass spectrometry method, most of the Eugenol content was 48.93%, followed by the Caryophyllene 20.78%, Methylis salicylas 14.96%. Through MIC, MBC and the experimental results showed that clove oil had antibacterial activity of Proteus, and with the increase of clove oil concentration, the inhibitory effect was better. When it reached a certain concentration, it can showed bactericidal effect that the MIC and MBC were 0.312 5%, 0.625 0%. The clove oil also had inhibitory effect on Streptococcus mutans biofilm with the concentration depedence. When the concentration of clove oil was 2.500 0%, the average fluorescence intensity (live/dead bacteria) was 0.082 ± 0.007, the proportion of living bacteria decreased obviously, and biofilm disappeared. Conclusions The clove oil not only has inhibitory effect on Streptococcus mutants, but also can inhibit and clear the biofilm formation of Streptococcus. Key words: Clove oil; Biofilm; Bacteria

Functional characterization of the eugenol synthase gene (RcEGS1) in rose

The floral volatile compound eugenol is an important constituent in many aromatic plants, being a floral attractant for pollinators as well as having antimicrobial activity. Rose flowers emit eugenol and its derivatives. We recently reported a eugenol synthase gene (RcEGS1) (JQ522949) that was present in petals of R. chinensis cv. Old Blush. RcEGS1 has its highest expression levels in the petals compared to other tissues; it has higher transcript levels at the developmental blooming stage and lower levels at budding and senescence stages. Here, we overexpressed the RcEGS1 protein in Escherichia coli, and showed by Western-blot analysis that its expression was mainly detected in stamens and petals at the flower opening stage. RcEGS1 was principally localized in the upper and lower epidermal layers, which are the major sites of scent emission in roses. Furthermore, we demonstrated that down-regulation of RcEGS1 expression in flowers by virus-induced gene silencing led to a reduction of the relative content of eugenol. We suggested that RcEGS1 was responsible for eugenol biosynthesis in roses.

Characteristics of eugenol loaded chitosan-tripolyphosphate particles as affected by initial content of eugenol and their in-vitro release characteristic

The aim of this research was to determine encapsulation efficiency, loading capacity and controlled release of eugenol loaded chitosan-tpp products which prepared by coaservation method. The characteristic of eugenol-loaded chitosan showed that %EE and % LC increased by increasing the initial eugenol content. The optimum of %EE (72.63%) and %LC (43.96%) were obtained at the ratio of chitosan to eugenol of 1:1.5. The FTIR spectrum showed the characteristic peaks of eugenol appearing on spectrum of eugenol encapsulated and blue-shift in the hydroxyl band from 3425.58 cm-1 in chitosan-tpp to 3417.86 cm-1 and 3394.72 cm-1 in eugenol loaded chitosan-tpp indicating that eugenol was successfully encapsulated. The surface morphologies of freeze-dried particles with the optimum %EE showed that more surface roughness and porosity than plain particles. Furthermore, the in vitro release of particles with minimum and optimum %EE were also investigated in acid (Simulated Gastric Fluid) and base (Simulated Intestinal Fluid) medium at ambient temperature.

Syzygium aromaticum : A subtropical tree scrutinized for the genetic differentiation among the altitudes of Nilgiris

Clove (Syzygium aromaticum) is one of the most economically significant oil producing trees in India, whose importance is mainly due to its medicinal features. Unfortunately, importing rate of the spice is increasing which would be an economic drawback for the country. Therefore, information regarding genetic diversity and the oil quality of the tree is necessary as a part to increase the cultivation. Thus the present work was conducted to identify the genetic difference between the collected samples among different altitudes of Nilgiris in comparison to the extracted oil quality to select high quality oil yielding variety for cropping. RAPD and GC-MS techniques were used for analyzing genetic diversity and oil extraction respectively. The phylogeny constructed revealed that the samples S1& S6 are genetically more similar, whereas S2 is far distinct from all other samples. S3 shared a close relation with S1 & S6. Samples S4, S5 were separated out but are within the same node. The eugenol percentage of the samples S1, S2, S3, S4, S5 and S6 are 42.96%, 68.78%, 55.95%, 71.17%, 74.04% and 73.47% respectively. From the collected data it is inferred that though the samples shared genetic similarity there observed a considerable difference in the eugenol content. This information needs to be correlated with the cultivation practices and the soil nutrition based on which commercial cropping could be done to improvize the clove sector of the country.

PHYTOCHEMICAL ANALYSIS OF SOME SELECTED VARIETIES OF PIPER BETLE L.

Objective: To Analyse the major phytochemical components in selected varieties of P. betle leaves using GC-MS.Methods: P. betle leaves were shade dried and pulverized to powder in a mechanical grinder. The powder was successively extracted with ethanol (40-60 °C). The extracts were concentrated under reduced pressure in a rotary evaporator. The ethanolic extracts of the plant leaves were used for GC-MS analysis.Results: In the present study ten chemicals were identified and compared from the P. betle varieties, the major component being Eugenol, which is responsible for the flavour and aroma of the leaves.Conclusion: Variety Karpoori possesses the highest content of Eugenol and could be used as a promising variety in the pharmaceutical industry. The components of essential oil can also serve as a determinant to distinguish different varieties of betle vine cultivars.

A greener approach towards isolating clove oil from buds of Syzygium aromaticum using microwave radiation

The selection of suitable technique and apposite operating conditions are the key attributes for efficient extraction of phytochemicals from plant materials. Microwave assisted extraction (MAE) was incorporated to isolate essential oil from clove buds and the parameters were optimized using the grey based Taguchi method to maximize the yield of clove oil (Y, % w/w), eugenol content (y, % w/w) and bacterial inhibition. The results of single response analysis and multi-response analysis were compared. It was possible to achieve the best results (Y: 13.11%, w/w; y: 11.93%, w/w; zone of inhibition (ZOI): 24.80 mm) using the optimized conditions of multi-response study viz., amount of plant material − 30 g; volume of water − 200 mL; power − 600 W and extraction time − 30 min. The wastes generated during extraction were explored for their utilization as a by-product or raw material for other processes. Moreover, the performance of MAE was compared with that of hydrodistillation in terms of processing conditions, responses, energy and environmental impact. It was possible to reduce water usage, extraction time, energy requirement and CO2 emission while achieving the improved quantity and quality of essential oil along-with a higher ZOI using the green MAE technique.

Estimation of Genetic Variability, Heritability and Genetic Advance for Essential Oil Yield and Related Traits in Genus Ocimum

Assessing variability is fundamental to identify the most important traits in Ocimum improvement program. The objective of the present study was to estimate variability, heritability and genetic advance based on twelve morphological characters of Ocimum. The experiment was conducted in 2015/16 main cropping season at one location by using randomized complete block design with three replications. The results revealed highly significant differences (P<0.01) among genotypes for all characters considered. The phenotypic coefficient of variation (PCV) for all character was higher than genotypic coefficient of variation (GCV). The estimation of broad sense heritability ĥ2(BS)% was observed to be lower than those of broad sense heritability ĥ2(BS)% for all characters. Genetics advance was recorded as maximum for eugenol content (96.65). Highest heritability in broad sense ĥ2(BS)% recorded for days to maturity (99.46). On the basis of study accessions G-4, G-7, G-9, G-11, G-18 and G-25 were identified high oil of better quality. These accessions may be exploited for commercial cultivation.

Effect of Methyl Jasmonate and Mannitol Application on Growth and Eugenol Content in Callus Cultures of Carnation

A protocol for callus induction from leaves of carnation is established. MS fortified with 2.0 mg /l 2,4‐D and 0.5 mg/l Kn was found to be the appropriate medium for producing callus cultures. Methyl jasmonate at 20 μM concentration showed the highest growth rate (98.21 mg/day) while mannitol concentrations at 2% gave the peak growth rate (97.1 mg/day).The highest content of eugenol (14.65 μg/100 g calli) was recorded with medium containing 20 μM methyl jasmonate followed by 8.16 μg/100 g calli with 10 μM methyl jasmonate. In presence of 0.5% mannitol, maximum eugenol content (18.00 μg/100 g calli) was obtained.Plant Tissue Cult. &amp; Biotech. 27(2): 227-240, 2017 (December)