Developed HPTLC Method Research Articles

DEVELOPMENT AND VALIDATION OF STABILITY INDICATING HPTLC METHOD FOR PIMAVANSERIN TARTRATE

Objective: The aim of current research work was to investigate degradation behavior of Pimavanserin tartrate upon exposure to stress conditions recommended by ICH Q1A (R2) and Q1B guidelines. Methods: Chromatographic separation was achieved on Merck’s TLC aluminum plates pre-coated with silica gel G 60 F254 as stationary phase and Methanol: Chloroform (2:8 v/v) as mobile phase. Densitometry scanning was carried out at 224 nm. Results: The retardation factor (Rf) was observed to be 0.56±0.02. Pimavanserin tartrate showed degradation in all stress conditions, but no degradation product was found in any stress condition. Peak purity was found to be 0.999 indicating no interference by degradation products to drug peak. The developed HPTLC method was successfully validated as per ICH Q2 (R1) guideline. Method was found to be linear within the range of 400-2000 ng/band with correlation coefficient R2= 0.9982. % RSD for intra-day and inter-day precision were found to be 1.35 and 1.78 % and % recovery was found to be in range 98-102 %. LOD and LOQ were found to be 17.58 ng/band and 53.27 ng/band respectively. Conclusion: A simple, economic stability indicating high performance thin layer chromatography method has been developed and validated for Pimavanserin tartrate. It is used for the treatment of delusions and hallucinations in Parkinson’s disease.

TLC-MS-Bioautographic identification of antityrosinase compounds and preparation of a topical gel formulation from bioactive fraction of RSM optimized alcoholic extract of Rubia cordifolia L. stem.

Rubia cordifolia L., Rubiaceae, is globally reported to treat skin-related problems. The study aimed to assess the antityrosinase potential of RC and the development of gel formulation. AutoDock vina (version V.1.2.0) program package was used for molecular docking to check for the binding affinity of ligands with protein. Response surface methodology (RSM) software was used to optimize extraction parameters for an alcoholic extract of Rubia cordifolia (ARC). The developed HPTLC method for the quantification of purpurin in ARC was validated as per the ICH guidelines, also the bioautographic study for evaluation of antityrosinase effects was performed, anthraquinone enriched fraction (AEF)-loaded gel formulation was developed and so evaluated which could be used to reduce skin pigmentation. Purpurin showed optimum binding affinity (-7.4 kcal/mol) with the molecular target (tyrosinase) when compared to that of standard kojic acid (-5.3 kcal/mol). Quantification of purpurin in ARC, optimized by RSM software was validated and physiologically significant results were observed for the antityrosinase potential of AEF, along with TLC-MS-bioautographic identification for antityrosinase compounds: purpurin (m/z 256.21) and ellagic acid (m/z 302.19). Evaluation of AEF-loaded gel formulation by in vitro and ex vivo permeation studies was performed. ARC extraction parameters optimized by RSM, and bioautographic study helped identify antityrosinase compounds. The development of gel formulation could be a cost-effective option for the treatment of depigmentation in the future.

Standardization and Quality Control Analysis of Marichadi tailam: An Ayurvedic Formulation

The Ayurvedic medicines are having great therapeutic application due to their natural holistic way of treatment of the disease. To ensure the effectiveness of the medicine standardization of the drug essential. Ayurvedic tailam is an important group of formulations used for treatment of various types of diseases. The principle of using tailam is better absorption of active ingredients by skin when applied through a lipophilic vehicle like vegetable oils. Marichadi Tailam is a herbal oil used for treatment of skin diseases such as eczema and dermatitis. In the present study, an attempt was made to develop and validate a HPTLC method for quantitative determination piperine in the Marichadi coconut oil and its standardization. The HTPLC separation was performed on precoated silica gel 60 F254 aluminum plate (10 × 10cm, 250μm thickness) using toluene:ethyl acetate:diethyl amine (7:2:1) as a mobile phase. The detection and quantification was performed at 340nm. The formulation contains 2.35% of piperine. Linearity studies indicated piperine in the linear ranges, while recovery studies revealed 99.32% (w/w) of piperine. The developed HPTLC method resolved and quantified piperine effectively, so it could be used as a simple reliable method for QC of polyherbal formulations.

HPTLC Fingerprint of an ayurvedic combination- Panchavalkala

Background: Panchavalkala is one of the most versatile ayurvedic combination of plants known for being used to reduce infections, inflammations and in wound healing and having a wide spectrum of medicinal uses. It has been formulated into various formulations like- Panchavalkala choorna, Lepa, Kashaya, kwatha, ointment, gel hand wash, decoction etc. It is It includes a group of five drugs- Ficus benghalensis L, Ficus racemosa L., Ficus religiosa L., Ficus lacor Buch. Ham, Thespesia populnea. WHO guidelines emphasises the need for modern and sophisticated modern instrumental method like HPTLC to standardize herbal materials. Aim and objective: To establish fingerprint profile of Panchavalkala combination and its individual drugs using HPTLC technique. Method: TLC method was developed for aqueous extract of individual drugs of Panchavalkala and Panchavalkala combination using solvent system- Toulene: ethyl acetate: methanol: formic acid (4:4:1:0.1). HPTLC fingerprinting was developed by scanning the chromatogram at 254, 366, 280 nm and at 250nm post derivatization using vanillin sulphuric acid. Results: The chromatograms were analyzed for number of peaks and common peaks in aqueous extract of individual drugs and Panchavalkala combination. HPTLC fingerprinting of all the individual extracts was established to identify them. The presence of common peaks found in individual extracts was confirmed in Panchavalkala combination, indicating that the developed HPTLC method can be successfully used to identify and standardize the combination. Conclusion: It can be concluded that HPTLC fingerprint analysis of Panchavalkala combination and its individual drugs can be used for Standardisation.

Marker Based Standardization of Herbal Sunscreen Formulation by Using Validated High-Performance Thin Layer Chromatography Method

Plant based products are worldwide used and trusted for the care and cure of the health. Herbal sunscreen formulations are gaining popularity for their effectiveness and are devoid of side effects. The present investigation is an effort made to develop herbal sunscreen cream containing methanolic extracts of different plants and its standardization for the presence of bioactive compounds. The leaves of Cymbopogon citratus (Stapf), fruit peel of Punica granatum (Linn), flowers of Butea monosperma (Lam.) and leaves of Neolamarckia cadamba (Roxb.) were selected for the preparation of sunscreen cream. The standardization of herbal formulations is very important to determine its quality based on the concentration of their active phytoconstituents. The herbal cream F-5 (2 % w/w) imparted its sun-protective and antioxidant property. It shows free radicals scavenging activity due the presence of flavonoids and phenolic compounds. High performance thin layer chromatography method is used to determine the quality and quantity of the sun protective phytoconstituents present in the product. The method was validated according to ICH guidelines for the estimation butrin (BT), isobutrin (IBT), quercetin (QC), apigenin (API), chlorogenic acid (CA) and gallic acid (GA) using the optimized solvent systems. The estimation of bioactive markers was carried out on silica gel precoated thin layer chromatography plates with 60F254 as the stationary phase and Camag TC scanner III for densitometric scanning. The average Rf values for the markers were found to be 0.46 for BT, 0.57 for IBT, 0.50 for QC, 0.57 for API, 0.66 for CA and 0.42 for GA. The developed HPTLC method was linear with correlation coefficient 0.999 for BT, 0.998 for IBT, QC, API and 0.9966 for CA and 0.9989 for GA. Limit of detection (LOD) and limit of quantification (LOQ) were recorded. The developed analytical method for quantitative determination of phytoconstituents was found efficient, simple, accurate, and validated.

Quantitative estimation of betulin in different plant parts of Dilleniapentagyna Roxb. through validated HPTLC method

A new, simple and accurate high-performance liquid chromatographic (HPTLC) method was developed for estimation of betulin (BE) in fruits, leaves, root bark and stem bark of Dillenia pentagyna , a RET species of Madhya Pradesh state of India. HPTLC aluminum plates precoated with 0.2 mm layer of silica gel 60F 254 were used to separate BE from the plant samples using the mobile phase, petroleum ether: ethyl acetate: acetonitrile (8: 2: 0.1, v/v/v). The anisaldehyde - sulphuric acid reagent was used to derivatize the plate followed by heating at 105°C on CAMAG HPTLC hot plate for 5 minutes. The peak corresponding to reference compound BE and that of the extract appeared at the same R F 0.21. Image of the plate was captured and HPTLC scan was performed densitometrically at ƛ max = 580 nm under max tungsten lamp in CAMAG TLC Scanner4 equipped with win CATS software. The developed HPTLC method was validated as per norms of the International Conference on Harmonization (ICH) for linearity, sensitivity, precision, accuracy and specificity. The BE content in fruit, leaf, root bark and stem bark samples of D. pentagyna was found as 0.101 ± 0.03%, 0.091 ± 0.09, 0.368 ± 0.07 and 0.440 ± 0.11% respectively.

A Study on HPTLC Quantification for Quality Control of Myricetin a Nutraceutical from Different Plant Parts of Myrica esculenta Linn

The present study was performed to report the quantification of myricetin present in different extracts (i.e. ethanol, methanol, petroleum ether, chloroform and aqueous extract) of root, leaf, bark, stem and seed of Myrica esculenta using TLC densitometric method. Densitometric scanning was performed at 366 nm for the determination and quantitation of standard compounds. The compact spots were achieved at Rf 0.67 corresponding to myricetin. The content of myricetin present in different solvent extracts of M. esculenta was 0.2159 ± 0.002% w/w, 0.3139 ± 0.003% w/w, 0.0184 ± 0.004% w/w, 0.0139 ± 0.003% w/w and 0.2120 ± 0.005% w/w in ethanolic, methanolic, petroleum ether, chloroform and aqueous extract of M. esculenta stem, stem, bark, leaf and leaf plant parts, respectively. The highest myricetin content was found in the order of methanolic stem extract>ethanolic stem extract>aqueous leaf extract>petroleum ether bark extract>chloroform leaf extract of M. esculenta. The developed HPTLC method was a precise, simple, convenient and accurate for screening of large number of active compounds present in various solvent extracts. Thus, the study concluded that the developed HPTLC method for quantification of myricetin can be used for a routine quality check and analysis of any formulation and composition containing M. esculenta.

Development and Validation of a Stability-Indicating HPTLC Method for the Estimation of Eszopiclone in Pharmaceutical Dosage Forms

Objective: A simple, sensitive, selective, precise repeatable and stability-indicating high-performance thin layer chromatographic method was developed and validated for Eszopiclone in bulk drug and in formulation. Method: Silica gel 60 F-254, TLC precoated aluminium plates was used as the stationary phase for analyzing Eszopiclone and its degradation products, using mobile phase consisting toluene: ethyl acetate: methanol (6: 4: 2 v/v/v). Result: This mobile phase gave compact spots for Eszopiclone with Rf value of 0.52 ± 0.02. Eszopiclone was exposed to hydrolysis, oxidation, neutral and photolytic conditions for conducting stress degradation study. The peak of Eszopiclone and the degradation product was well resolved from each other with a significantly different Rf value. Densitometric estimation of Eszopiclone was performed at 304nm. A good linear plot was obtained in the concentration range of 150-300ng/spot. The method was validated for precision, accuracy (recovery) and robustness study. The limit of detection (LOD) and limit of quantitation (LOQ) was found to be 130ng/spot and 150ng/spot, respectively. Conclusion: The developed HPTLC method can separate Eszopiclone from its degradation products, hence stability studies can be performed using this method.

High performance thin-layer chromatography and in vitro cytotoxic studies on ethanol extract of Matricaria chamomilla L (Asteraceae) flowers

Purpose: To develop a high performance thin-layer chromatography (HPTLC procedure for quantitation of apigenin in ethanol extract of Matricaria chamomilla (Babunaj) flowers, and to evaluate the extract for in vitro cytotoxic effect on MCF-7 cell lines.
 Methods: Quantification of apigenin was carried out using a CAMAG TLC system. A combination of toluene, ethyl acetate and formic acid (4.5:3.5:0.2 v/v/v) was used as mobile phase, with densitometry detection at 336 nm. The HPTLC procedure was subjected to validation as per ICH guidelines. The cytotoxicity of the extract was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay.
 Results: A sharp apigenin band at Rf of 0.51 was obtained, and the content of apigenin in the extract was 0.062 % w/w. The detection limit (LOD) and quantification limit (LOQ) were 0.19 and 0.57 ng/band, respectively. MTT assay results indicate that M. chamomilla was cytotoxic to Michigan Cancer Foundation-7 (MCF-7) cells, with half-maximal concentration (IC50) of 74 µg/mL.
 Conclusion: The developed HPTLC method is linear, precise, accurate and specific for the determination of apigenin. M. chamomilla exerts cytotoxic effect on MCF-7 cell line via induction of apoptosis.

Taxonomy, Pharmacognosy and Phytochemical Characteristics to Identify the Authentic and Substitute Botanical Sources Used as Agnimantha

The roots of Clerodendrum phlomidis L.f. is known to be used as Agnimantha in Ayurvedic system of medicine. Whereas literature emerged from various texts reports other species of Premna as Agnimantha that leads to controversies in usage of authentic drug source. Hence the study aims to develop pharmacognosy characteristics to identify the authentic and substitute sources of Agnimantha. The Ayurvedic verses prescribed for Agnimantha compliance with Clerodendrum phlomidis, Premna serratifolia and Premna mollissima. Whereas the vertically fissured stem found prominent in Clerodendrum phlomidis. Likewise cork region is composed of homogenous cells with grouped and scattered calcium oxalate crystals, without oils. Whereas heterogeneous cells with oils and scattered calcium oxalate crystals are found in Premna serratifolia and Premna mollissima. Similarly phytosterols found unique in Clerodendrum phlomidis roots. The developed HPTLC method serves as tool to identify and distinguish the authentic from substitute and adulterants. Enthrallingly quantitative HPLC analysis revealed the presence of verbascoside in all sources of Agnimantha and comparatively more in substitutes. Hence the study concludes that roots of Clerodendrum phlomidis is authentic source of Agnimantha, where Premna serratifolia and Premna mollissima forms the equivalent substitutes.

Detection and Quantification of Anti-oxidant markers like Rutin, Catechin, Quercetin, Gallic acid, Ellagic acid, Ferulic acid, Vitexin and Mangiferin in Herbal raw materials available in market belongs to Rutaceae family by HPTLC Technique

The primary aim of the study is to detect the flavonoids, phenolic acids and xanthones in three commercial herbal raw materials namely Aegle marmelos, Citrus aurantifolia and Murraya koenigii obtained from Rutaceae family used in daily domestic needs to confirm the presence of mentioned common secondary metabolites in herbal raw materials. Results of the study clearly revealed that these raw materials from rutaceae family contains flavonoids, phenolic acids and xanthones. The developed HPTLC method can be employed for the routine investigations of flavonoids, phenolic acids and xanthones in herbal raw materials. Aegle marmelos, Citrus aurantifolia and Murraya koenigii was ascertain the presence of Rutin and shows the 0.072%,0.005%0.002% respectively. Ellagic acid was originate to be not more than 0.004% each of Aegle marmelos and Citrus aurantifolia. Quercetin was found to 0.034% .062% 0.021% respectively in all test three extracts. Vitexin was found to be 0.019% in Citrus aurantifolia. Murraya koenigii revels presence of 0.002%, 0.001% of mangiferin and catechin respectively

Validated HPTLC Method to Estimate Silymarin in Marketed Herbal Formulation

Objectives: The objective of present research work is to develop simple, rapid and accurate high performance thin layer chromatographic method for quantitative estimation of silymarin in marketed hepatoprotective formulation. Methods: The separation was performed on silica gel 60 F254 precoated TLC aluminum plates and toluene: ethyl acetate: formic acid: methanol as mobile phase. Determination and quantitation were performed by densitometric scanning at 288 nm. Validation of method is carried out as per International Conference of Harmonization (ICH) guidelines. Results: Accuracy, precision and recovery were all within the required limits. Conclusion: In conclusion, the developed HPTLC method is precise and accurate for implementation in routine quality control. The selected marketed hepatoprotective formulation contains the sufficient amount of the active ingredient which is required for its therapeutic efficiency.

Development of High Performance Thin Layer Chromatography Method for Quantitation of Gallic Acid in Normacid Syrup: A Poly Herbal Formulation

Normacid syrup consisiting of 17 ingredients, used in the treatment of hyperacidity and gastritis. Gallic acid is one of the phytoconstituent present in Normacid syrup- a poly herbal formulation. In the present study an attempt has been made to develop a HPTLC method for quantitative estimation of gallic acid in dried fruits used in formulations. This HPTLC method was found to be reproducible, accurate, precise and detect gallic acid at nanogram level. The developed HPTLC method would be an important tool in the quality control method of poly herbal formulation. 
 Keywords: HPTLC, Polyherbal, Gallic acid, Gastritis

DEVELOPMENT OF HIGH PERFORMANCE THIN LAYER CHROMATOGRAPHY METHOD FOR QUANTITATION OF ?-SITOSTEROL IN NORMACID SYRUP: A POLY HERBAL FORMULATION

Normacid syrup consisiting of 17 ingredients, used in the treatment of hyperacidity and gastritis. ?- Sitosterol is one of the phytoconstituent present in Normacid syrup- a poly herbal formulation. In the present study an attempt has been made to develop a HPTLC method for quantitative estimation of ?- Sitosterol in dried plant parts used in formulations. This HPTLC method was found to be reproducible, accurate, precise and detect ?- Sitosterol at nanogram level. The developed HPTLC method would be an important tool in the quality control method of poly herbal formulation. 
 Keywords: HPTLC, Polyherbal, ?- Sitosterol, Gastritis

Simultaneous Quantification of Pharmacologically Active Alkaloid Metabolites Colchicine and Gloriosine in Gloriosa Superba L. collected from Western Ghats (India) and Adjoining Areas for the Identification of Elite Chemotype(s).

Gloriosa superba is a valuable Ayurvedic medicinal plant and is in high demand in the world market for its colchicine content, which is used to treat gout. The study aims (1) to record the metabolic variations in major bioactive metabolites, colchicine and gloriosine, in the natural populations of G. superba from Western Ghats and adjoining areas in India and (2) to develop HPTLC protocol for the identification of elite chemotypes of species and regulation of quality raw material, extract, and finished material. Simultaneous quantification of colchicine and gloriosine in 22 natural populations through validated HPTLC as per ICH guidelines. Colchicine and gloriosine were identified at Rf 0.51 ± 0.03 and 0.41 ± 0.05 and the content varied from 0.021 to 0.86% and 0.003 to 0.198%. The method was found linear at a concentration range of 0.1-0.7 µg/spot, and LOD (3.3 σ/S) and LOQ (10 σ/S) was 0.71 and 2.16 µg/spot. The method was precise in the concentration range of 100-300 ng/spot, with 98.29% and 101.12% recovery (% RSD) for colchicine and gloriosine. Subsequently, four elite chemotypes were identified based on cluster analysis of metabolite content. The developed HPTLC method is linear, accurate, precise, and robust for simultaneous quantification of colchicine and gloriosine metabolite(s). Intraspecific metabolic variation was significant among the collected population, leading to the identification of four elite chemotypes. Colchicine is an industrially viable metabolite and is therefore quintessential to the development of an economical and analytical method to regulate the quality of raw material, extract, and finished products.

Simultaneous Analytical Efficiency Evaluation Using an HPTLC Method for the Analysis of Syringic Acid and Vanillic Acid and Their Anti-Oxidant Capacity from Methanol Extract of Ricinus communis L. and Euphorbia hirta L.

The well-known medicinal plants Ricinus communis L. and Euphorbia hirta L. have been traditionally used to treat a variety of ailments in the India and other countries. In the present research, a simple, rapid, reliable, and accessible HPTLC technique has been established for concurrent quantification of phenolic compounds (syringic acid and vanillic acid) and evaluation of their anti-oxidant capacity. The chromatographic separation was conceded on pre-coated silica gel plates 60 F254 as the stationary phase. The appropriate mobile phase combination of toluene-ethyl acetate-formic acid (7:2.5:0.5, v/v/v) were developed to expand the plates which separated components according to the marker compounds. Further, the anti-oxidant potential of methanol extract of R. communis (MERC) and E. hirta (MERH) were also assessed with 1,1-diphenyl-2-picrylhydrazyl (DPPH) by using UV spectrophotometry. Densitometric scanning was performed with a Camag V scanner and measured at two different wave lengths, 272 and 318 nm. The marker compounds were practically resolved with RF 0.5 ± 0.04 for syringic acid and 0.6 ± 0.06 for vanillic acid. The results obtained in the study of anti-oxidant activity of MERC and MERH showed significant free radical scavenging capacity against DPPH-generated free radicals. The developed HPTLC method was validated for accuracy, linearity, precision, and specificity. Both the extracts revealed considerable antioxidant activity. The reported existing phenolic and flavonoids compounds are responsible for antioxidant activity of plant extracts.

Development and validation of high performance thin layer chromatographic method for the determination of voglibose in bulk and their formulations

To develop a simple, fast, specific, precise and accurate High Performance Thin Layer Chromatographic method (HPTLC) for the determination of Voglibose in bulk and their dosage forms. The chromatographic separation was achieved on precoated silica gel 60F254 aluminum plates using a combination of acetonitrile: methanol: ammonia (15:4:0.1 % V/V/V) as mobile phase and densitometric evaluation of spots was carried out at 284 nm using Camag TLC scanner III with CATS 1.3.4 version software. The experimental parameters like band size of the chamber saturation time, spot application, slit width, solvent front migration, etc. were studied critically and evolved optimized conditions. The drug was well resolved satisfactorily with Rf value 0.66±0.03. The repeatability and accuracy of the optimized method were ascertained by evaluating various validation parameters like linearity (100 to 450 ng/spot), precision (intra-day % RSD 0.21 to 0.74, inter-day % RSD 0.21 to 0.29), accuracy (99.8% to 101.2%, % RSD below 1%), and specificity according to ICH guidelines. The limits of detection and quantification were 40 ng/spot and 100 ng/spot respectively. The developed HPTLC method was faster and cost effective quantitative control for routine analysis of Voglibose in bulk and its formulations.

Impact of seasonal variation on ‘daidzein’ accumulation in callus and in vivo parts of Pueraria tuberosa (Willd.) DC

Daidzein-natural isoflavones found in Pueraria tuberosa (Willd.) DC. (Fabaceae) which is a potent medicinal plant. It acts as signal carriers and responds to the pathogenic attacks and reduces pain during menopause, osteoporosis and also antidiabetic in nature. In the present work, in vitro callus development from tuber explants during both rainy and summer seasons was conducted. Further, a simple unambiguous and rapid high-performance thin-layer chromatography method was established for quantitative estimation of daidzein in callus and in vivo parts to assess the impact of seasonal change on valuable phytochemical accumulation. Maximum callusing (90%) was obtained on MS medium fortified with a combination of 6N-Benzylamino purine (2 mg/l) and 2,4-dichlorophenoxyacetic acid (2 mg/l) during the summer season from the tuber explant. During the HPTLC method validation, the linearity range obtained was 100–1000 ng/spot with a regression value (r) value of 0.99845. All the parts were found to contain a significant amount of daidzein. The maximum daidzein (2112.567±0.35 ng/g) content was obtained from a young tuber bark followed by callus (171.903±0.33 ng/g) during the summer season compared to rainy season parts and callus. Thus, it can be concluded that in vitro callus is an alternative source of daidzein without destroying the natural plant and the developed HPTLC method could be used for quality control analysis and recommended for daidzein quantification for different herbal formulation and drug preparation.

Development and validation of a high-performance thin-layer chromatographic method for the quantitative analysis of vitexin in Passiflora foetida herbal formulations

The aim of this study is the development of validated HPTLC method for the quantification of vitexin from Passiflora foetida commercial herbal formulations. The developed method was validated, in accordance with ICH guidelines for precision, accuracy, specificity and robustness. The plate was developed using ethyl acetate:methanol:water:formic acid 30:4:2:1(%, v/v/v/v) on 20 × 10 cm glass coated silica gel 60 F254 plates and the developed plate was scanned and quantified densitometrically at λ = 340 nm. Linear regression analysis revealed a good linear relationship between peak area and amount of vitexin in the range of 100–700 ng/spot. The amount of vitexin in nine commercial herbal formulations was successfully quantified by the developed HPTLC method. The developed and validated high performance thin layer chromatographic method offers a new sensitive and reliable tool for quantification of vitexinin in various herbal formulations containing Passiflora foetida.

Antitick potential and chemical variability among Ageratum conyzoides L. germplasms collected from Eastern and Western Ghats of India

ABSTRACTAgeratum conyzoides L. is a common weed in many tropical and subtropical countries including India. The present study aimed to evaluate the antitick potential using Adult Immersion Test (AIT) and variability of its phytoconstituents with the developed HPTLC method in germplasms collected from Eastern and Western Ghats of India. Percent tick mortality at 8% concentration ranged from 10.0 ± 0.2 to 85.0 ± 2.9. Precocene I content ranged from 0.0004% to 0.072%, Precocene II content varied from 0.001% to 0.035%, and Caryophyllene oxide content was found to range from 0.0001% to 0.0023% in crude drug. Precocene II was positively correlated (coefficient = 0.656) with tick mortality (%) in the germplasm collected from Western Ghats. However, in Eastern Ghat samples, Precocene I was positively correlated (coefficient = 0.550) with per cent tick mortality. There is significant variation in marker compound content as well as antitick activity of the extracts; however, a direct correlation among marker compounds and antitick activity was not observed. This indicates there may be some other phytomolecules responsible for antitick activity of the extract. This study will lead to a potential source of quality raw material for the industry in developing herbal acaricide.