Bioassay-guided detection, identification and assessment of antibacterial and anti-inflammatory compounds from olive tree flower extracts by high-performance thin-layer chromatography linked to spectroscopy.

Olive trees are one of the most widely cultivated fruit trees in the world. The chemical compositions and biological activities of olive tree fruit and leaves have been extensively researched for their nutritional and health-promoting properties. In contrast, limited data have been reported on olive flowers. The present study aimed to analyse bioactive compounds in olive flower extracts and the effect of fermentation-assisted extraction on phenolic content and antioxidant activity. High-performance thin-layer chromatography (HPTLC) hyphenated with the bioassay-guided detection and spectroscopic identification of bioactive compounds was used for the analysis. Enzymatic and bacterial in situ bioassays were used to detect COX-1 enzyme inhibition and antibacterial activity. Multiple zones of antibacterial activity and one zone of COX-1 inhibition were detected in both, non-fermented and fermented, extracts. A newly developed HPTLC-based experimental protocol was used to measure the high-maximal inhibitory concentrations (IC50) for the assessment of the relative potency of the extracts in inhibiting COX-1 enzyme and antibacterial activity. Strong antibacterial activities detected in zones 4 and 7 were significantly higher in comparison to ampicillin, as confirmed by low IC50 values (IC50 = 57-58 µg in zone 4 and IC50 = 157-167 µg in zone 7) compared to the ampicillin IC50 value (IC50 = 495 µg). The COX-1 inhibition by the extract (IC50 = 76-98 µg) was also strong compared to that of salicylic acid (IC50 = 557 µg). By comparing the locations of the bands to coeluted standards, compounds from detected bioactive bands were tentatively identified. The eluates from bioactive HPTLC zones were further analysed by FTIR NMR, and LC-MS spectroscopy. Multiple zones of antibacterial activity were associated with the presence of triterpenoid acids, while COX-1 inhibition was related to the presence of long-chain fatty acids.

Bioassay-Guided Assessment of Antioxidative, Anti-Inflammatory and Antimicrobial Activities of Extracts from Medicinal Plants via High-Performance Thin-Layer Chromatography.

Natural products and their analogues have contributed significantly to treatment options, especially for anti-inflammatory and infectious diseases. Thus, the primary objective of this work was to compare the bioactivity profiles of selected medicinal plants that are historically used in folk medicine to treat inflammation and infections in the body. Chemical HPTLC fingerprinting was used to assess antioxidant, phenolic and flavonoid content, while bioassay-guided HPTLC was used to detect compounds with the highest antibacterial and anti-inflammatory activities. The results of this study showed that green tea leaf, walnut leaf, St. John's wort herb, wild thyme herb, European goldenrod herb, chamomile flower, and immortelle flower extracts were strong radical scavengers. Green tea and nettle extracts were the most active extracts against E. coli, while calendula flower extract showed significant potency against S. aureus. Furthermore, green tea, greater celandine, and fumitory extracts exhibited pronounced potential in suppressing COX-1 activity. The bioactive compounds from the green tea extract, as the most bioactive, were isolated by preparative thin-layer chromatography and characterized with their FTIR spectra. Although earlier studies have related green tea's anti-inflammatory properties to the presence of catechins, particularly epigallocatechin-3-gallate, the FTIR spectrum of the compound from the most intense bioactive zone showed the strongest anti-inflammatory activity can be attributed to amino acids and heterocyclic compounds. As expected, antibacterial activity in extracts was related to fatty acids and monoglycerides.

Evaluation of bioactive compounds from Ficus carica L. leaf extracts via high-performance thin-layer chromatography combined with effect-directed analysis.

This study compares different solvent systems with the use of spontaneous fermentation on the phytochemical composition of leaf extracts from a locally grown white variety of common fig (Ficus carica Linn.). The aim was to detect and identify bioactive compounds that are responsible for acetylcholinesterase (AChE), α-amylase and cyclooxygenase-1 (COX-1) enzyme inhibition, and compounds that exhibit antimicrobial activity. Bioactive zones in chromatograms were detected by combining High-performance thin-layer chromatography (HPTLC) with enzymatic and biological assays. A new experimental protocol for measuring the relative half-maximum inhibitory concentration (IC50) was designed to evaluate the potency of the extracts compared to the potency of known inhibitors. Although the IC50 of the fig leaf extract for α-amylase and AChE inhibition were significantly higher when compared to IC50 for acarbose and donepezil, the COX-1 inhibition by the extract (IC50 = 627 µg) was comparable to that of salicylic acid (IC50 = 557 µg), and antimicrobial activity of the extract (IC50 = 375-511 µg) was similar to ampicillin (IC50 = 495 µg). Four chromatographic zones exhibited bioactivity. Compounds from detected bioactive bands were provisionally identified by comparing the band positions to coeluted standards, and by Fourier transform infrared (FTIR) spectra from eluted zones. Flash chromatography was used to separate selected extract into fractions and isolate fractions that are rich in bioactive compounds for further characterisation with nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS) analysis. The main constituents identified were umbelliferon (zone 1), furocoumarins psoralen and bergapten (zone 2), different fatty acids (zone 3 and 4), and pentacyclic triterpenoids (calotropenyl acetate or lupeol) and stigmasterol (zone 4).

Effect directed analysis of bioactive compounds in leaf extracts from two Salvia species by High-performance thin-layer chromatography.

Extracts of two Salvia species, Salvia apiana (white sage) and Salvia officinalis (common sage) were screened for phytoconstituents with the ability to act as antidiabetic, cognitive enhancing, or antimicrobial agents, by hyphenation of high-performance thin-layer chromatography with enzymatic and microbial effect directed assays. Two bioactive zones with α-amylase inhibition (zone 1 and zone 2), 3 zones for acetylcholinesterase inhibition (zones 3, 4 and 5), and two zones for antimicrobial activity (zones 4 and 5) were detected. The compounds from the five bioactive zones were initially identified by coelution with standards and comparing the RF values of standards to the bioautograms. Identity was confirmed with ATR-FTIR spectra of the isolated compounds from the bioactive zones. A significantly higher α-amylase and acetylcholinesterase inhibition of S. apiana leaf extract was associated with a higher flavonoid and diterpenoid content. Fermented S. officinalis extract exhibited a significantly higher ability to inhibit α-amylase compared to other non-fermented extracts from this species, due to increased extraction of flavonoids. The ATR-FTIR spectra of 2 zones with α-amylase inhibition, indicated that flavonoids and phenolic acids were responsible for α-amylase inhibition. Multiple zones of acetylcholinesterase inhibition were related to the presence of phenolic abietane diterpenoids and triterpenoid acids. The presence of abietane diterpenoids and triterpenoid acids was also found responsible for the mild antimicrobial activity. Flash chromatography was used to isolate sufficient amounts of bioactive compounds for further characterisation via NMR and MS spectroscopy. Five compounds were assigned to the zones where bioactivity was observed: cirsimaritin (zone 1), a caffeic acid polymer (zone 2), 16-hydroxyrosmanol (zone 3), 16-hydroxycarnosic acid (zone 4), oleanolic and ursolic acids (zone 5).

The bioprofiling of antibacterials in olive leaf extracts via thin layer chromatography-effect directed analysis (TLC-EDA).

In this study, effect-directed analysis (EDA) (i.e. TLC hyphenated with an in situ MTT (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide) antimicrobial assay), was used for screening and identification of antimicrobials in olive leaf extract. EDA detected that the same compounds exhibited significant antimicrobial activity against bacterial species of the genera Enterococcus (E. faecalis), Escherichia (E. coli), Streptococcus (S. mutans) and Staphylococcus (S. aureus). Flash chromatography-fractionation was used to isolate antimicrobial compounds in olive leaf extract. The active compounds were identified as maslinic acid and oleanolic acid by comparing RF values of the detected active bands with the standard reference materials, with identity confirmed with NMR and ATR-FTIR spectroscopy. Maslinic and oleanolic acids were tested on the E. faecalis strain (which displayed the highest sensitivity in the MTT assay) to determine their inhibiting concentration 50% (IC50) and minimum bactericidal concentrations.

HPTLC and ATR/FTIR Characterization of Antioxidants in Different Rosemary Extracts.

The effect of spontaneous fermentation by lactic acid bacteria on the extraction yield of bioactive compounds and antioxidant activity from rosemary leaf extracts was investigated using high-performance thin-layer chromatography (HPTLC). Brining and spontaneous fermentation with lactic acid bacteria more than doubled extraction of polyphenolics and antioxidants from the rosemary leaves. The results show that lactic acid fermentation enhances antioxidant activity in extracts by increasing the total phenolic content but does not increase extraction of phytosterols. Increased extraction of phenolic oxidants during fermentation assisted extraction, results from the in situ generated natural eutectic solvent from the plant sample. ATR-FTIR spectra from the bioactive bands suggests that this increased antioxidant activity is associated with increased extraction of rosmarinic acid, depolymerised lignin, abietane diterpenoids and 15-hydroxy-7-oxodehydroabietic acid.

The effect of extractive lacto-fermentation on the bioactivity and natural products content of Pittosporum angustifolium (gumbi gumbi) extracts.

Pittosporum angustifolium, known as gumbi gumbi, is a native Australian plant, which has traditionally been used as an Aboriginal medicine. This study investigates the effect of different solvents and extractive fermentation on the content and natural products composition of Pittosporum angustifolium extracts, and compares their antioxidant activity, in vitro α-amylase inhibition, and anti-inflammatory properties. Anti-inflammatory activity of the extracts was determined by measuring the inhibition of nitric oxide (NO) production. Extracts were characterised with FTIR-ATR spectroscopy, and screened for antioxidant activities and α-amylase inhibitory activity via High-performance thin-layer chromatography (HPTLC)-Effect-directed analysis (EDA) with direct bioautography. HPTLC combined with chemical derivatization and bioassays was used for EDA screening. The results show that lactic acid fermentation of gumbi gumbi leaves boosts the antioxidant activity in extracts by increasing the total phenolic content, but does not affect (increase or decrease) α-amylase inhibitory activity or nitrogen scavenging/anti-inflammatory activity. Analysis of the ATR-FTIR spectra from the band at RF = 0.85 that inhibits α-amylase, suggests that fatty acid esters are responsible for the enzyme inhibition; both saturated fatty acid esters in unfermented extracts and unsaturated fatty acid esters in fermented extracts. The ATR-FTIR spectra of the polyphenolics in fermented extracts (RF = 0.15-0.20) suggests the presence of soluble lignin fragments (i.e. lignins depolymerized into monomers and oligomers during the fermentation process).

Characterisation of α-amylase inhibitors in marigold plants via bioassay-guided high-performance thin-layer chromatography and attenuated total reflectance-Fourier transform infrared spectroscopy.

A high-performance thin-layer chromatography with microchemical derivatization and bioassay guided detection was used for bioanalytical profiling of selected marigold plant extracts. Anisaldehyde/sulfuric acid reagent and thymol/sulfuric acid reagent were used to visualize separated components on the chromatograms. Antioxidant activity and α-amylase inhibition were assessed with 2 bioassays, DPPH assay to detect free radical scavengers and starch-iodineassay method to detect compounds that inhibit α-amylase. The highest antioxidant activity of 10.12 µg of gallic acid equivalents (GAE) per 20 µL of extract was measured in extract from Tagetes flowers and the lowest in the extract from Calendula leaves with 5.10 µg of GAE. Multiple zones of α-amylase inhibition were detected. A detailed analysis of the ATR-FTIR spectra from the bands at RF = 0.24 suggest that faradiol esters and saturated fatty acids esters, palmitic acid, myristic acid, and lauric acid are responsible for α-amylase inhibition, unsaturated fatty acids for the band at RF = 0.51 and phytoecdysteroids for the band at RF = 0.53.

A new integrated HPTLC - ATR/FTIR approach in marine algae bioprofiling.

The aim of this study was to evaluate marine algae extracts in terms of their anti-inflammatory activity using a combination of chromatographic separation and chemical detection with subsequent infrared vibrational spectroscopy identification. Extraction parameters, chemical fingerprint, and the activity levels were considered for the method optimization. High-performance thin-layer chromatography (HPTLC) combined with microchemical derivatization, was used to separate and detect bioactive compounds with antioxidant activity and anti-inflammatory activities, and to detect different classes of terpenoids. Infrared attenuated total reflectance (ATR) spectral analysis of the bands with bioactive compounds, identified sulfated polysaccharides to be responsible for the anti-inflammatory activity in extracts of brown algae Carpoglossum confluens and Phyllospora comosa. Steroids as unique antioxidants with significant free radical scavenging activities were observed in extracts of brown algae Cystophora platylobium, Cystophora retorta, Carpoglossum confluens and Phyllospora comosa. HPTLC combined with biochemical assays and FTIR-ATR spectrometry was demonstrated to be a straightforward strategy for bioprofiling marine algae extracts.

High-performance thin-layer chromatography linked with (bio)assays and FTIR-ATR spectroscopy as a method for discovery and quantification of bioactive components in native Australian plants.

Many native Australian plants have a long history of use as medicinal and culinary herbs and some are considered to be equivalents to the Mediterranean herbs. However, while therapeutic properties of Mediterranean herbs are well documented, there is limited information on the medicinal use of the Australian native herbs. Extracts of five native Australian plants were characterised with FTIR-ATR spectroscopy in the fingerprint region and screened for enzyme inhibitory and antioxidant activities via effect-directed analysis (EDA) based on bioautography. High-performance thin-layer chromatography (HPTLC) coupled with microchemical and biochemical derivatization assays was used for EDA screening. Detected compounds with biological activities were identified via FTIR-ATR spectroscopy. All herbs showed antioxidant activity with lemon myrtle being the most active. The α-amylase inhibition, observed in native thyme, sea parsley and native bush was associated with the presence of phenolic acids, chlorogenic acid and caffeic acid. The investigation of botanicals by a fast, hyphenated HPTLC method, has allowed an effect-directed high-throughput screening, fast characterization of complex mixtures and detection of biologically active phytochemicals (bioprofiling).

Bioassay-guided identification of α-amylase inhibitors in herbal extracts.

This study focuses on the health benefits of several fresh herbs that are commonly used in the Mediterranean diet. Antioxidant activity, phytosterol content and α-amylase inhibitory activity of fresh basil, lavender, oregano, rosemary, sage, and thyme are analyzed and compared. High-performance thin-layer chromatography (HPTLC) combined with effect directed analysis was used to detect and quantify biological active compounds on chromatograms. The highest antioxidant activity was measured in the extract from oregano leaf, while the highest terpenoid content was in basil leaf extract. All extracts except lavender leaf and lavender flower extracts showed α-amylase inhibition. The same compound at hRF = 68 in basil, oregano, rosemary, sage, and thyme extracts was responsible for α-amylase inhibition. Combined with effect-directed assays and attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, hyphenated HPTLC allowed a fast characterization of the active compound. ATR spectral analysis of this band tentatively identified oleanolic acid (or its derivative) to be responsible for the α-amylase inhibition.

Models for skin and brain penetration of major components from essential oils used in aromatherapy for dementia patients.

Aromatherapy with essential oils (EOs) has been linked to improvement of cognitive function in patients with dementia. In order to act systemically, active EO components must be absorbed through the skin, enter the systemic circulation, and cross the blood brain barrier (BBB). Thus, the aim of this work was to develop quantitative structure activity relationships (QSARs), to predict skin and blood barrier penetrative abilities of 119 terpenoids from EOs used in aromatherapy. The first model was based on experimentally measured skin permeability for 162 molecules, and the second model on BBB permeability for 138 molecules. Each molecule was encoded with 63 calculated molecular descriptors and an artificial neural network was used to correlate molecular descriptors to permeabilities. Developed QSAR models confirm that EOs components penetrate through the skin and across the BBB. Some well-known descriptors, such as log P (lipophilicity), molecular size and shape, dominated the QSAR model for BBB permeability. Compounds with the highest predicted BBB penetration were hydrocarbon terpenes with the smallest molecular size and highest lipophilicity. Thus, molecular size is a limiting factor for penetration. Compounds with the highest skin permeability have slightly higher molecular size, high lipophilicity and low polarity. Our work shows that a major disadvantage of novel multitarget compounds developed for the treatment of Alzheimer's disease is the size of molecules, which cause problems in their delivery to the brain. Therefore, there is a need for smaller compounds, which possess more desirable physicochemical properties and pharmacokinetics, in addition to targeted biological effects.Communicated by Ramaswamy H. Sarma.

High-Performance Thin-Layer Chromatography Hyphenated with Microchemical and Biochemical Derivatizations in Bioactivity Profiling of Marine Species.

Marine organisms produce an array of biologically active natural products, many of which have unique structures that have not been found in terrestrial organisms. Hence, marine algae provide a unique source of bioactive compounds. The present study investigated 19 marine algae and one seagrass collected from Torquay beach, Victoria, Australia. High-performance thin-layer chromatography (HPTLC) hyphenated with microchemical (DPPH•, p-anisaldehyde, and Fast Blue B) and biochemical (α-amylase and acetylcholine esterase (AChE) enzymatic) derivatizations was used to evaluate antioxidant activity, presence of phytosterols and phenolic lipids, α-amylase and AChE inhibitory activities of extract components. Significant α-amylase and AChE inhibitory activities were observed in samples 2, 6, 8 and 10. Antioxidant activities in the samples were found to be correlated to phytosterol content (R² = 0.78), but was not found to be related to either α-amylase or AChE inhibitory activities. α-Amylase inhibitory activities were correlated to AChE inhibition (R² = 0.77) and attributed to the phytosterol content, based on the similar peak position in the chromatograms with the ß-sitosterol chromatogram. Samples 1, 8, and especially sample 20, were found to contain phenolic lipids (alkyl resorcinol derivatives) with significant antioxidant activities. The results suggest that these marine species have a significant number of bioactive compounds that warrant further investigation.

HPTLC - Bioautographic methods for selective detection of the antioxidant and α-amylase inhibitory activity in plant extracts.

A high-performance thin-layer chromatography (HPTLC) method was developed for quantification of α-amylase inhibitory activity and stigmasterol content in ant plant extracts. An improved HPTLC method for the determination of total free radical scavenging activity in samples using DPPH• is also reported. For quantification of α-amylase inhibitory activity, the developed HPTLC plate is dipped into an α-amylase solution, and the bioautogram is then incubated at 25 °C for 30 min under humid conditions. For visualization of enzyme inhibitory activity, the starch test with an iodine indicator solution is used. The blue zone observed comes from the starch-iodine complex formed from starch that was not hydrolyzed by the amylase due to enzyme inhibition by the compound(s) present in the sample. The area of the blue zones was used to compare and quantify relative α-amylase inhibitory activity in different extracts. Location of the blue zones (hRF) on the plate was used to detect compounds that are responsible for the α-amylase inhibitory activity. Relative α-amylase activity was not related to the antioxidant activity, but was highly correlated with the stigmasterol content in the sample extracts (R = 0.95). Therefore, plant sterols present in the extracts might be responsible for α-amylase inhibitory activities in the extracts. •The developed method for quantification of α-amylase inhibitory activity provides an efficient and effective tool that can be used to screen, detect and quantify α-amylase inhibitory activity in plant extracts.•The proposed protocol is easy to run, involves minimal sample preparation, with multiple samples able to be analyzed in parallel on the same chromatographic plate, in a short time.•There were significant differences in α-amylase inhibitory activity, stigmasterol content, and total free radical scavenging activity between methanol, ethanol, dichloromethane, and ethyl acetate ant plant extracts.

High-performance thin-layer chromatography HPTLC-direct bioautography as a method of choice for alpha-amylase and antioxidant activity evaluation in marine algae.

High-Performance Thin-layer chromatography (HPTLC) combined with DPPH free radical method and α-amylase bioassay was used to compare antioxidant and antidiabetic activities in ethanol and ethyl acetate extracts from 10 marine macroalgae species (3 Chlorophyta, 4 Phaeophyta and 3 Rhodophyta) from Blue Lagoon beach (Malaysia). Samples were also evaluated for their phenolic and stigmasterol content. On average, higher antioxidant activity was observed in the ethyl acetate extracts (55.1mg/100g gallic acid equivalents (GAE) compared to 35.0mg/100g GAE) while, as expected, phenolic content was higher in ethanol extracts (330.5mg/100g GAE compared to 289.5mg/100g GAE). Amounts of fucoxanthin, stigmasterol and α-amylase inhibitory activities were higher in ethyl acetate extracts. Higher enzyme inhibition is therefore related to higher concentrations of triterpenes and phytosterols (Note: these compounds are more soluble in ethyl acetate). Ethyl acetate extracts from Caulerpa racemosa and Padina minor, had the highest α-amylase inhibitory activity, and also showed moderately high antioxidant activities, stigmasterol content and polyphenolic content. Caulerpa racemose, being green algae, does not contain fucoxanthin, while Padina minor, being brown algae, contains high amounts of fucoxanthin. Therefore, it is very unlikely that fucoxanthin contributes to α-amylase inhibitory activity as previously reported.

Probing into the Molecular Requirements for Antioxidant Activity in Plant Phenolic Compounds Utilizing a Combined Strategy of PCA and ANN.

AIM AND OBJECTIVE: This study investigates molecular structural requirements that are responsible for the antioxidant activity in phenolic compounds. METHOD: Antioxidant activity of compounds was determined with a 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical assay. Principal component analysis (PCA) was used to classify phenolic antioxidants according to the key molecular features that contribute to their antioxidant activity. Artificial neutral networks (ANNs) was used to develop a predictive QSAR model. RESULTS: Both models agreed that structural characteristics of phenolic compounds responsible for the antioxidant activity include: (1) number and position of alcohol groups on the aromatic ring; (2) molecular size; (3) flexibility/bulkiness; and (4) water solubility. PCA has classified data into phenolic acids and flavonoids, suggesting two distinct mechanisms of action. ANN has confirmed different mechanisms of action for flavonoids and polyphenolic acids, i.e. breaking of free radical chain reactions by donation of a hydrogen atom to neutralise a free radical and the chelating ability of polyphenolic acids. CONCLUSION: Although two phenolic acids may have the same relative polarity, their different functional groups may drastically change the nature of their interactions with free radicals, and their antioxidant activity.

Assessment of antioxidant activity in Victorian marine algal extracts using high performance thin-layer chromatography and multivariate analysis.

The aim of this study was to develop and validate a rapid and simple high performance thin layer chromatographic (HPTLC) method to screen for antioxidant activity in algal samples. 16 algal species were collected from local Victorian beaches. Fucoxanthin, one of the most abundant marine carotenoids was quantified directly from the HPTLC plates before derivatization, while derivatization either with 2,2-diphenyl-1-picrylhydrazyl (DPPH) or ferric chloride (FeCl3) was used to analyze antioxidants in marine algae, based on their ability to scavenge non biological stable free radical (DPPH) or to chelate iron ions. Principal component analysis of obtained HPTLC fingerprints has classified algae species into 5 groups according to their chemical/antioxidant profiles. The investigated brown algae samples were found to be rich in non-and moderate-polar compounds and phenolic compounds with antioxidant activity. Most of the phenolic iron chelators also have shown free radical scavenging activity. Strong positive and significant correlations between total phenolic content and DPPH radical scavenging activity showed that, phenolic compounds, including flavonoids are the main contributors of antioxidant activity in these species. The results suggest that certain brown algae possess significantly higher antioxidant potential when compared to red or green algae and could be considered for future applications in medicine, dietary supplements, cosmetics or food industries. Cystophora monilifera extract was found to have the highest antioxidant concentration, followed by Zonaria angustata, Cystophora pectinate, Codium fragile, and Cystophora pectinata. Fucoxanthin was found mainly in the brown algae species. The proposed methods provide an edge in terms of screening for antioxidants and quantification of antioxidant constituents in complex mixtures. The current application also demonstrates flexibility and versatility of a standard HPTLC system in the drug discovery. Proposed methods could be used for the bioassay-guided isolation of unknown natural antioxidants and subsequent identification if combined with spectroscopic identification.

Rapid evaluation and comparison of natural products and antioxidant activity in calendula, feverfew, and German chamomile extracts.

The present study describes a simple high performance thin layer chromatographic (HPTLC) method for the simultaneous quantification of apigenin, chamazulene, bisabolol and the use of DPPH free radical as a post-chromatographic derivatization agent to compare the free radical scavenging activities of these components in leaf and flower head extracts from feverfew, German chamomile and marigold from the Asteraceae family. Feverfew (Tanacetum parthenium) leaves have been traditionally used in the treatment of migraine with parthenolide being the main bioactive compound. However, due to similar flowers, feverfew is sometimes mistaken for the German chamomile (Matricaria recutita). Bisabolol and chamazulene are the main components in chamomile essential oil. Marigold (Calendula officinalis) was included in the study for comparison, as it belongs to the same family. Parthenolide was found to be present in all leaf extracts but was not detected in calendula flower extract. Chamazulene and bisabolol were found to be present in higher concentrations in chamomile and Calendula flowers. Apigenin was detected and quantified only in chamomile extracts (highest concentration in flower head extracts). Antioxidant activity in sample extracts was compared by superimposing the chromatograms obtained after post-chromatographic derivatization with DPPH and post-chromatographic derivatization with anisaldehyde. It was found that extracts from chamomile flower heads and leaves have the most prominent antioxidant activity, with bisabolol and chamazulene being the most effective antioxidants.

High performance thin layer chromatography (HPTLC) and high performance liquid chromatography (HPLC) for the qualitative and quantitative analysis of Calendula officinalis-advantages and limitations.

Chromatography techniques such as HPTLC and HPLC are commonly used to produce a chemical fingerprint of a plant to allow identification and quantify the main constituents within the plant. The aims of this study were to compare HPTLC and HPLC, for qualitative and quantitative analysis of the major constituents of Calendula officinalis and to investigate the effect of different extraction techniques on the C. officinalis extract composition from different parts of the plant. The results found HPTLC to be effective for qualitative analysis, however, HPLC was found to be more accurate for quantitative analysis. A combination of the two methods may be useful in a quality control setting as it would allow rapid qualitative analysis of herbal material while maintaining accurate quantification of extract composition.