Investigation of the composition and stability of the essential oils of origanum vulgare ssp.vulgare L. andO. vulgare ssp.hirtum (Link) letswaart

Summary The essential oils ofOriganum vulgare L. ssp.hirtum (Link) letswaart andOriganum vulgare L ssp.vulgare (Fam. Lamiaceae), cultivated in Hungary, have been studied by GC and GC-MS and the qualitative and quantitative chemical composition of the essential oils in the two species have been compared.O. vulgare ssp.hirtum oil was found to contain carvacrol (76.4%), γ-terpinene (6.6%), thymol (0.23%), andp-cymene (4.7%) as the main constituents whereas the major compounds inO. vulgare ssp.vulgare oil werep-cymene (22.3%), caryophyllene oxide (10.2%), sabinene (7.9%), γ-terpinene (5.1%), thymol (0.34%), and spathulenol (4.8%). The stability of content and composition of the oils during the flowering period (economically beneficial period) were observed. The effect of long-term storage on the composition of the oil was also investigated for both the crude and distilled oil ofOriganum vulgare ssp.vulgare. Presented at Balaton Symposium '01 on High-Performance Separation Methods, Siófok, Hungary, September 2–4, 2001     

Determination of phenolic compounds by MALDI–TOF and essential oil composition by GC–MS during three development stages of Origanum majorana L.

This study aimed to investigate the effect of the maturation process of sweet marjoram (Origanum majorana L.) on essential oil composition, the phenolic profile of ethanolic extract and their antioxidant capacities. The essential oil composition was studied at three stages of maturity by GC–MS. Thirty compounds were detected representing 100% of the total essential oil. p‐Menth‐1‐en‐4‐ol was the major compound (37.15–76.94%) followed by cyclohexanol‐3,3,5 trimethyl (5.41–15.99%) and α‐terpineol (0.94–11.34%). During the maturation process, an accumulation of oxygenated monoterpenes was observed. The phenolic composition was studied using matrix‐assisted laser desorption/ionization time of flight. The analysis showed the presence of short flavonoid monomers at all stages of maturation. The antioxidant capacity of ethanolic extracts and essential oils was evaluated using the DPPH assay, iron chelating power and reducing power assay. The highest phenolic content and antioxidant capacity were found at flowering stage. These findings on essential oil composition, phenolic profile and antioxidant capacity of O. majorana at three different stages of development provide more information on how these secondary metabolites are accumulated.     

Comparison of the oxidative stability of soybean and sunflower oils enriched with herbal plant extracts

The present study was conducted to determine and compare the oxidative stability of soybean and sunflower oils using differential scanning calorimetry (DSC). These edible oils were enriched with marjoram (Origanum majorana L.), thyme (Thymus vulgaris L.), and oregano (Origanum vulgare L.) extracts at three different concentrations and synthetic antioxidant (BHA). The fatty acid composition of studied oils was determined by gas chromatography mass spectrometry to evaluate the content of unsaturated fatty acids that are sensitive to oxidation process. Oil samples were heated in the DSC at different heating rates (4.0, 7.5, 10.0, 12.5, and 15.0 °C min^?1) and oxidation kinetic parameters (activation energy, pre-exponential factor, and oxidation rate constant) were calculated. The results showed that the oxidative stability of sunflower oil samples enriched with oregano extracts and soybean oil supplemented with thyme extracts was improved compared to samples without the addition of herbal plant extracts and the synthetic antioxidant.     

Optimization of carvacrol,rosmarinic, oleanolic and ursolic acid extraction from oregano herbs (Origanum onites L., Origanum vulgare spp. hirtum and Origanum vulgare L.)

The aim of our study was to increase the extraction efficiency of carvacrol, rosmarinic, oleanolic and ursolic acid from the different species of oregano herbs (Origanum onites L., Origanum vulgare spp. hirtum and Origanum vulgare L.). Various extraction methods (ultrasound-assisted, heat-reflux, continuous stirring, maceration, percolation) and extraction conditions (different solvent, material:solvent ratio, extraction temperature, extraction time) were used, and the active substances were determined by HPLC. The lowest content of carvacrol, rosmarinic, oleanolic and ursolic acid was obtained by percolation. During heat-reflux extraction, the content of active substances depended on the solvent used: ethanol/non-aqueous solvent (glycerol or propylene glycol) mixture was more effective compared with ethanol alone. The results showed that for each species of oregano the most optimal extraction method should be selected to maximize the content of biologically active substances in the extracts.     

Determination of phenolic compounds in medicinal plants from the Lamiaceae family

A procedure for the determination of Phenolic compounds in extracts from the medicinal plants of the Lamiaceae family—garden sage (Salvia officinalis L.), creeping thyme (Thymus serpyllum L.), wild marjoram (Origanum vulgare L.), and common balm (Melissa officinalis L.)—obtained under different extraction conditions was developed. The identification of the extracted compounds was performed and their qualitative and quantitative composition was established by HPLC with diode array and mass-spectrometric detection with consideration for the obtained characteristics of the standard samples of individual components. The test samples of medicinal herbs contained caffeic acid (0.19–0.62 mg/g) and rosmaric acid (4–23 mg/g); the highest rosmaric acid content (23 mg/g) was found in wild marjoram, and the lowest content (4 mg/g), in creeping thyme. The extracts of wild marjoram contained the greatest amounts of Phenolic compounds; rosmaric acid and luteolin-7-O-β-D-glucuronide were major components, whereas protocatechuic, 3-O-caffeoylquinic, and caffeic acids were minor components.     

Simultaneous determination of phenolic compounds and triterpenic acids in oregano growing wild in Greece by 31P NMR spectroscopy

³¹P nuclear magnetic resonance (NMR) spectroscopy was used to detect and quantify simultaneously a large number of phenolic compounds and the two triterpenic acids, ursolic acid and oleanolic acid, extracted from two oregano species Origanum onites and Origanum vulgare ssp. Hirtum using two different organic solvents ethanol and ethyl acetate. This analytical method is based on the derivatization of the hydroxyl and carboxyl groups of these compounds with the phosphorous reagent 2‐chloro‐4,4,5,5‐tetramethyl‐1,3,2‐dioxa phospholane and the identification of the phosphitylated compounds on the basis of the ³¹P chemical shifts. Unambiguous assignment of the ³¹P NMR chemical shifts of the dihydroxy‐ and polyhydroxy‐phenols in oregano species as well as those of the triterpenic acids was achieved upon comparison with the chemical shifts of model compounds assigned by using two‐dimensional NMR techniques. Furthermore, the integration of the appropriate signals of the hydroxyl derivatives in the corresponding ³¹P NMR spectra and the use of the phosphitylated cyclohexanol as an internal standard allowed the quantification of these compounds. The validity of this technique for quantitative measurements was thoroughly examined. Copyright © 2012 John Wiley & Sons, Ltd.     

Insecticidal properties of essential oils against Tribolium castaneum (Herbst) and their inhibitory effects on acetylcholinesterase and adenosine triphosphatases

Essential oils from 20 Egyptian plants were obtained by using hydrodistillation. The chemical composition of the isolated oils was identified by gas chromatograph/mass spectrometer. Fumigant and contact toxicities of the essential oils were evaluated against the adults of Tribolium castaneum. In fumigation assays, the oil of Origanum vulgare (LC₅₀ = 9.97 mg/L air) displayed the highest toxicity towards the adults of T. castaneum. In contact assays, the oils of Artemisia monosperma (LC₅₀ = 0.07 mg/cm²) and O. vulgare (LC₅₀ = 0.07 mg/cm²) were the most potent toxicants against the adults of T. castaneum. Biochemical studies showed that the tested oils caused pronounced inhibition of acetylcholinesterase (AChE) and adenosine triphosphatases (ATPases) isolated from the larvae of T. castaneum. The oil Cupressus macrocarpa (IC₅₀ = 12.3 mg/L) was the most potent inhibitor of AChE, while the oil of Calistemon viminals (IC₅₀ = 4.4 mg/L) was the most potent inhibitor of ATPases.     

Agronomic,metabolomic and lipidomic characterisation of Sicilian Origanum vulgare (L.) ecotypes

Although Origanum vulgare (L.) has been deeply analysed at phytochemical level, poor knowledge is available regarding non-volatile compounds such as lipids. The aim of this work was to characterise five wild Sicilian Origanum ecotypes from an agronomic, metabolomic and lipidomic perspective. Serradifalco presented higher dry weight and inflorescences/plant than the others while Favara had a significantly higher number of branches per plant and more extensive flowered stratum. Metabolomic analysis, performed with LC-MS-TOF, allowed a preliminary characterisation of the non-volatile metabolome of the five oregano ecotypes Origanum vulgare ssp. hirtum. Twenty-five metabolites were identified belonging to organic acids, amino acids, lysophosphatidylcholines, carnithines, nucleic bases and lysophosphatidylethanolamines. Lipidomic analysis identified 115 polar plant membrane glycerolipid species. Thirteen of them were differentially present in the two chosen ecotypes. The role of these metabolites in plant physiology from a qualitative and pharmacological point of view was discussed.     

An Improved On-Line HPLC-DPPH Method for the Screening of Free Radical Scavenging Compounds in Water Extracts of Lamiaceae Plants

An on-line HPLC-1,1-diphenyl-2-picrylhydrazyl (DPPH*) method has been improved for the detection of polar and nonpolar radical scavenging compounds from complex plant extracts. Eight water extracts were prepared from steam-distilled essential oil-extracted Lamiaceae plants (Origanum vulgare L., O. Onites L., O. Minutiflorum O. Schwartz et P. H. Davis, O. Syriacum L., Satureja cuneifolia Ten., Thymbra spicata L., Coridothymus capitatus (L.) Reichb. f., Majorana hortensis Moench). After the components within each extract had been separated by reverse phase chromatography using 10% to 100% methanol with 2% acetic acid as a mobile phase, analytes capable of scavenging a citric acid-sodium citrate buffered methanolic DPPH* solution were detected by post-column derivatization at 517 nm. The HPLC-DPPH* on-line method was applied to the qualitative and quantitative analysis of these Lamiaceae plant extracts. There was a strong correlation between the scavenging (negative) peak area and the concentration of the radical scavenging reference substances used. The radical scavenging compounds within the extracts were determined as benzoic acid and hydroxycinnamic acid derivatives, flavonoids and diterpenoids according to their retention time and UV spectral data. Rosmarinic acid and carnosic acid were identified as the dominant radical scavengers in these extracts by this method.     

Polyphenol composition and antioxidant activity of selected medicinal herbs

The aim of this study was to characterize aqueous and alcoholic extracts [30%, 50% and 70% (w/v)] obtained from medicinal herbs (Calendula officinalis, Hypericum perforatum, Galium verum, and Origanum vulgare) used in traditional medicine from our country. Samples were examined for total and individual content of phenolics and antioxidant activities. The highest content of total polyphenols (9.9 ± 0.02 mg gallic acid equivalents (GAE) L^–1 extract) and antioxidant activities expressed as Trolox Equivalent Antioxidant Capacity [307.51 TEAC mmol g^–1DW by the ABTS (2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) method and 20.90 TEAC mmol g^–1 DW by the DPPH (1,1-diphenyl-2-picrylhydrazyl) method] was found in Origanum vulgare (50%) extract. Polyphenolic compounds were quantified using RP-HPLC.     

Comparison of the origin and phenolic contents of Lycium ruthenicum Murr. by high‐performance liquid chromatography fingerprinting combined with quadrupole time‐of‐flight mass spectrometry and chemometrics

The fruits of Lycium ruthenicum Murr. have long been used in folk medicine. Nevertheless, detailed information related to its phenolic composition and its quality control remains scarce. In this study, a simple and reproducible method, based on high‐performance liquid chromatography combined with chemometrics, was developed to authenticate 18 samples of L. ruthenicum Murr. collected from different parts of China through fingerprint analysis. The main peaks were identified by quadrupole time‐of‐flight electrospray ionization mass spectrometry. Four phenolics were quantified, and the most abundant phenolic compound in almost all samples was kukoamine A. Hierarchical cluster analysis and principal component analysis were applied to classify these samples. Also, a total of 26 compounds, which were mainly phenolic compounds and anthocyanins, were identified or tentatively identified based on the available literature and standard references. Among these, 16 were reported for the first time in the extract. The results showed that there was no significant difference between L. ruthenicum fruits from different provinces in terms of chemical composition. Also, the fingerprint together with chemometric analyses and quadrupole time‐of‐flight electrospray ionization mass spectrometry are promising methods for evaluating the quality consistency, identification, and comprehensive evaluation of L. ruthenicum .     

Crude extract of Origanum vulgare L. induced cell death and suppressed MAPK and PI3/Akt signaling pathways in SW13 and H295R cell lines

Oregano (Origanum vulgare L.) is a common aromatic plant used in Mediterranean and Asian Regions for treating respiratory diseases, painful menstruation, rheumatoid arthritis, etc. Recently its role as an anticancer plant has been suggested, although oregano has been never evaluated into adrenocortical tumour cell models. This study analysed for the first time the anticancer effects of a crude extract of wild mountain oregano (Origanum vulgare L.) in SW13 and H295R cell lines. The crude extract was characterised by GC/MS and the toxic effects of oregano were first analysed by brine shrimp lethality assay. Our findings demonstrated that oregano decreased cell viability, survival, modified cell cycle and induced cell death (through necrotic process) and that the effects can be attributed to a blockade of MAPK and PI3 K/Akt pathways. These results suggest that oregano extract exerts anticancer activities in adrenocortical tumour cell lines, providing evidence for further research in higher models.     

Trace elements and essential oil composition in chemotypes of the aromatic plantOriganum vulgare

Trace elements, essential oil yield and its percentage composition were determined by neutron activation analysis, hydrodistillation and gas chromatography in two chemotypes ofOriganum vulgare L. cultivated in the same field. Statistical tests such as analysis of variance, correlation coefficient,t-test, and multiple correlation were applied. The results showed that the samples contain the highest recorded oil yield for aromatic plants. Also, there is a statistically significant difference between the chemotypes of the plant not only in the predominant compound but in a number of other components. Iron, chromium and scandium showed a negative significant correlation with carvacrol and a positive one with thymol. Europium shows characteristic correlations with chromium, cobalt, iron and scandium within each chemotype of the plant. These correlations could make probable a role of this element in the biosynthesis of the predominant compounds.     

Shelf Life Extension and Quality Improvement of Cucumber Slices Impregnated in Infusions of Edible Herbs

Abstract

Development of functional food through their enrichment with herbal extracts is gaining much attention from the food industry. The present study evaluates the benefits, such as the oxidative stability and quality retention, of the enrichment of cucumber (Cucumis sativus) with natural phenolic compounds through immersion into a variety of herbal infusions. After a preliminary experiment with gallic acid solutions, the beneficial effect of hypotonic solutions was observed and the optimal process conditions were defined. The second step of the study involved the immersion of cucumber in the herbal infusions for 2.5?h at 70?°C. The total phenolic content (TPC), antiradical and antioxidant activity of osmo-treated slices were determined to evaluate the extent of bioactive compounds’ impregnation. During the immersion, TPC, antioxidant and antiradical activity significantly increased irrespective of the type of the herbal infusion. The TPC varied greatly, ranging from 1.41 to 781.14?mg GAE/kg of cucumber. The highest total phenolics were found in cucumber treated with Origanum vulgare infusion, followed by Jasminum officinale and Mentha spicata infusions. The aforementioned osmo-treated slices were also highly appreciated by the sensory panel; therefore, for those best performing herbal infusions, the third phase of this work involved the study of quality degradation under subsequent storage of the osmo-treated slices vs. the untreated ones. The results regarding color, texture, and visual assessment demonstrated the superior quality retention of the osmo-treated samples that exhibited a shelf life extension ranging from two- to almost four-fold compared to the untreated tissue.     

Spectroscopy,Microscopy and Fluorescence Imaging of Origanum vulgare L. Basis for Nondestructive Quality Assessment

The organs of Origanum vulgare L. plant were examined by optical microscopy, scanning electron microscopy and autofluorescence imaging. The different organs were also studied spectroscopically. Fluorescence emission spectra were recorded for intact inflorescences, leaves and stems. Several fluorescence ratios (Blue/Red, Blue/Far‐red, Green/Red and Green/Far‐red), which varied depending on the considered organ of the plant, were derived. For leaves, a dependence of fluorescence spectra with water content was obtained as well. The intact samples were also analyzed by Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy. These spectra were transformed to the Remission function depending on the wavenumber and two absorption bands (811 and 1740 cm^?1), which displayed differences according to the plant organ sampled, were detected. These results were consistent with higher carvacrol content in inflorescences. The spectroscopic results were connected with the microscopic observation and with the presence of relevant nutraceutics contained in the plant. The optical indexes derived in this work may serve as potential indicators to be explored in the development of nondestructive methods for oregano quality assessment.     

Determination of the Free Radical Scavenging Activity of Lycium Extracts

Lycium species growing in Turkey have not so far been studied sufficiently. For this reason, non-polar and polar extracts obtained from the fruits of Lycium barbarum L. and L. Ruthenicum Murray (Solanaceae) were assessed both in vitro for their potential as free radical scavenger crude extracts and their phenolic composition. Fruits of Lycium species were sequentially extracted with petroleum ether, ethyl acetate, methanol, n-butanol, and water in a Soxhlet extractor. All the extracts were assessed for the scavenging of the nitrogen-centered free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH^.) by in vitro method. Furthermore, the composition of each extract was investigated both in terms of its Folin-Ciocalteau reactive components and its qualitative content. The phenolic compounds within the extracts were determined as benzoic acid and hydroxycinnamic acid derivatives, flavonoids, and anthocyanins according to their retention time and UV spectral data by HPLC-DAD system.     

Simultaneous determination of eight major steroids from Polyporus umbellatus by high‐performance liquid chromatography coupled with mass spectrometry detections

Polyporus umbellatus is a widely used diuretic herbal medicine. In this study, a high‐performance liquid chromatography coupled with atmospheric pressure chemical ionization–mass spectrometric detection (HPLC‐APCI‐MS) method was developed for qualitative and quantitative analysis of steroids, as well as for the quality control of Polyporus umbellatus. The selectivity, reproducibility and sensitivity were compared with HPLC with photodiode array detection and evaporative light scattering detection (ELSD). Selective ion monitoring in positive mode was used for qualitative and quantitative analysis of eight major components and β‐ecdysterone was used as the internal standard. Limits of detection and quantification fell in the ranges 7–21 and 18–63 ng/mL for the eight analytes with an injection of 10 µL samples, and all calibration curves showed good linear regression (r² > 0.9919) within the test range. The quantitative results demonstrated that samples from different localities showed different qualities. Advantages, in comparison with conventional HPLC–diode array detection and HPLC‐ELSD, are that reliable identification of target compounds could be achieved by accurate mass measurements along with characteristic retention time, and the great enhancement in selectivity and sensitivity allows identification and quantification of low levels of constituents in complex Polyporus umbellatus matrixes. Copyright © 2009 John Wiley & Sons, Ltd.     

Sub-inhibitory stress with essential oil affects enterotoxins production and essential oil susceptibility in Staphylococcus aureus

Fourteen wild strains of Staphylococcus aureus positive for gene sea were tested for enterotoxins production and the minimum inhibitory concentration of Leptospermum scoparium, Origanum majorana, Origanum vulgare, Satureja montana and Thymus vulgaris essential oils (EOs) were determined. After this trial, bacteria stressed with sub-inhibitory concentration of each EO were tested for enterotoxins production by an immunoenzymatic assay and resistance to the same EO. Oregano oil exhibited the highest antibacterial activity followed by manuka and thyme oils. After the exposure to a sub-inhibitory concentration of EOs, strains displayed an increased sensitivity in more than 95% of the cases. After treatment with oregano and marjoram EOs, few strains showed a modified enterotoxins production, while 43% of the strains were no longer able to produce enterotoxins after treatment with manuka EO. The results obtained in this study highlight that exposure to sub-inhibitory concentration of EO modifies strains enterotoxins production and EOs susceptibility profile.     

Chemical Characterization and Antibacterial Activity of Essential Oil of Medicinal Plants from Eastern Serbia

The objective of this study was to evaluate wild growing Satureja kitaibelii, Thymus serpyllum, Origanum vulgare, Achillea millefolium and Achillea clypeolata with respect to their essential oil (EO) content, composition and antimicrobial activity. The five species were collected at Mt. Rtanj and the village of Sesalac, Eastern Serbia. The main EO constituents of Lamiaceae plants were p-cymene (24.4%), geraniol (63.4%) and germacrene D (21.5%) in Satureja kitaibelii, Thymus serpyllum and Origanum vulgare ssp. vulgare, respectively. A. millefolium EO had multiple constituents with major ones being camphor (9.8%), caryophyllene oxide (6.5%), terpinen-4-ol (6.3%) and 1,8-cineole (5.6%), while the main EO constituents of A. clypeolata were 1,8-cineole (45.1%) and camphor (18.2%). Antimicrobial testing of the EO showed that Staphylococcus aureus (Gram-positive) was more sensitive to all of the tested EOs than Escherichia coli (Gram-negative). S. kitaibelii EO showed the highest antimicrobial activity against both tested bacterial strains. This is the first study to characterize the EO composition and antimicrobial activity of these five medicinal species from Eastern Serbia in comparison with comprehensive literature data. The results can be utilized by the perfumery, cosmetics, food and pharmaceutical industries, but also for healing purposes in self-medication.     

Quality evaluation of Salvia miltiorrhiza Bge. by ultra high performance liquid chromatography with photodiode array detection and chemical fingerprinting coupled with chemometric analysis

An ultra high performance liquid chromatography with photodiode array detection method is developed for the simultaneous quantitative determination of five water‐soluble compounds including danshensu, protocatechualdehyde, rosmarinic acid, salvianolic acid B, and salvianolic acid A in Salvia miltiorrhiza Bge. samples. Through method optimization, the five compounds all expressed good linearity (R² > 0.9990) in a wide concentration range together with satisfactory accuracy, precision, and stability. Moreover, through qualitative analysis of the chemical fingerprint combined with similarity analysis, hierarchical cluster analysis, principle component analysis, and partial least‐squares discriminate analysis, we determined that the 13 batches of Salvia miltiorrhiza Bge. were similar in internal quality and the differences resulted from various cultivation environments, recovery elements, and others. Seen from the results of hierarchical cluster analysis and principle component analysis, the classification of 13 batches was in accordance, and partial least‐squares discriminate analysis technique was more suitable than the principle component analysis model to provide a distinct classification of test samples on the basis of their different components. Moreover, a permutation test verified the rationality of partial least‐squares discriminate analysis and variable importance plot showed that peaks 37 and 38 were the most significant variables in distinguishing the Salvia miltiorrhiza Bge. samples. The idea of the quantitative and qualitative analysis of Salvia miltiorrhiza Bge. was convenient, sensitive, and comprehensive, which could be applied to evaluate the quality of more traditional Chinese medicines.