Search results for: hydro-distillation

Authors: Imen Blibech, Mohiedine Ksantini, Mohamed Bouaziz

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Prays oleae is a major insect of olive in the Mediterranean Region. In an effort to find effective and affordable ways of controlling this pest, larvicidal activity of essential oils from Tunisian Thymus capitatus were analyzed in comparison to Azadirachtin, a biologically active compound insecticide. The essential oils were extracted by hydrodistillation, and their chemical composition was determined by gas liquid-chromatography coupled with mass spectroscopy. The main components of chemical components were oxygenated monoterpenes (60.24%). The most abundant oxygenated monoterpenes were carvacrol (54.11%). Monoterpenes hydrocarbons were much more abundant and dominated by the o-cymene (16.68%). Both active compounds of Azadirachtin and Thymus capitatus oil extracts exhibited significant larvicidal activity against P. oleae with LC50 values 81.30 ppm and 52.49 ppm respectively. Dose-response relationships were established with almost 100% mortality when using the highest dose 100 ppm of T. capitatus oil extracts and 80 ppm of Azadirachtin. At the lowest dose (10 ppm), T. capitatus oil extracts and Azadirachtin caused 60% and 76% larval mortality in 48 hours respectively. The larval mortality rate greatly decreased with increases of the dilution of both oil extract compounds. Larval development duration appeared to be prolonged to about 12 days for larvae feeding on control diet. The maximum antifeedant activity was shown by both T. capitatus oil extract and Azadirachtin at LC90 values (47.5 and 50.1 ppm respectively). Tunisian T. capitatus oil extract used at low concentrations could be considered as eco-friendly promising insecticide similar to Azadirachtin that has significant potential for the biological control of P. oleae.

Keywords: Thymus capitatus, chemical composition, azadirachtin, larvicidal effects, antifeedant activity, Prays oleae

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Authors: Dalvir Kataria, Khushminder Kaur Chahal, Amit Kumar

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The carrot seed essential oil was obtained by hydrodistillation. Hexane, dichloromethane, and methanol solvents were used for extraction of carrot seed by Soxhlet extraction methods. The major and minor compounds identified in carrot seed essential oil were carotol (52.73), daucol (5.10), daucene (5.68), (E)-β-farnesene (5.40), β-cubebene (3.19), longifolenaldehyde (3.23), β-elimene (3.23), (E)-caryophyllene (1.22), β-bisabolene (2.95) etc. The chemical composition of hexane, dichloromethane, and methanol extracts was different. Carotol was the common compound present. Major compounds isolated were from the carrot seed essential oil by column chromatography. Chemical transformations of carotol (2) with mercuric acetate/sodium borohydride, dry hydrochloric acid gas, acetonitrile/sulfuric acid, selenium dioxide/t-butyl hydrogen peroxide, N-bromosuccinimide, hydrogen iodide, and phenol were carried out. The derivatives of carotol were designed to explore the significance of some structural modifications in relation to antioxidant activities. The structures of major compounds and derivatives were confirmed on the basis of FT-IR, 1HNMR and 13CNMR spectroscopy. Antioxidant activity of carrot seed essential oil, various extracts and isolated compounds were tested by in vitro models involving 2, 2-diphenyl-1-picrylhydrazyl (DPPH•), hydroxyl (OH•), nitric oxide (NO•), superoxide radical scavenging methods and ferric reducing antioxidant power assay (FRAP). Chemical transformations of major isolated compound carotol were carried out, and antioxidant activity of all compounds was undertaken. The major sesquiterpenoidcarotol isolated from carrot seed essential oil showed the highest antioxidant activity in all the methods. The methanol extract showed higher antioxidant potential as compared to carrot seed essential oil, hexane, and dichloromethane extracts.

Keywords: antioxidant, carotol, carrot, DPPH

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Authors: Nassim Belkacem, Amina Azzam, Dalila Haouchine, Kahina Bennacer, Samira Soufit

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Objective: To evaluate the antioxidant activity of the methanolic extract along with the antimicrobial activity of the essential oil of the aerial parts of Rosmarinus officinalis L. collected in the region of Bejaia (northern center of Algeria). Materials and methods: The polyphenols and flavonoids contents of the methanolic extract were measured. The antioxidant activity was evaluated using two methods: the ABTS method and DPPH assay. The antimicrobial activity was studied by the agar diffusion method against five bacterial strains (Three Gram positive strains and two Gram negative strains) and one fungus. Results: The total polyphenol and flavonoid content was about 43.8 mg gallic acid equivalent per gram (GA Eq/g) and 7.04 mg quercetin equivalent per gram (Q Eq/g), respectively. In the ABTS assay, the rosemary extract has shown an inhibition of 98.02% at the concentration of 500ug/ml with a half maximal inhibitory concentration value (IC50) of 194.92ug/ml. The results of DPPH assay have shown that the rosemary extract has an inhibition of 94.67 % with an IC50 value of 17.87ug/ml, which is lower than that of Butylhydroxyanisol (BHA) about 6.03ug/ml and ascorbic acid about 1.24μg/ml. The yield in essential oil of rosemary obtained by hydrodistillation was 1.42%. Based on the determination of the diameter of inhibition, different antimicrobial activity of the essential oil was revealed against the six tested microbes. Escherichia coli from the University Hospital (UH), Streptococcus aureus (UH) and Pseudomonas aeruginosa ATCC have a minimum inhibitory concentration value (MIC) of 62.5µl/ml. However, Bacillus sp (UH) and Staphylococcus aureus ATCC have an MIC value of 125μl/ml. The inhibition zone against Candida sp was about 24 mm. The aromatograms showed that the essential oil of rosemary exercises an antifungal activity more important than the antibacterial one.

Keywords: Rosmarinus officinalis L., maceration, essential oil, antioxidant, antimicrobial activity

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Authors: Elsa Makue Nguuffo, Esther Del Florence Moni Ndedi, Jacky Njiki Bikoï, Jean Paul Assam Assam, Maximilienne Ascension Nyegue

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Aims: The present study aims to evaluate the kinetic parameters of essential oils (EOs) and combinations fromDrypetes gossweileri Stem Bark, Ocimum gratissimum leaves, Cymbopogon citratusleaves after evaluation of their antibacterial activityonmultidrug-resistant strains ofShigella. Material and Methods:fiveclinical strains of Shigellaisolated from patients with diarrhoeaincluding Shigella flexneri, and 4 otherstrains of Shigella sppwere selected. Their antibiotic profile was established using agar test diffusion with seven antibiotics belonging to seven classes.EOs were extracted from each plant using hydrodistillation process. The activity of Ciprofloxacin®, OEs, and their combination formulatedinthe followingratios(w/w/w): C1: 1/1/1; C2: 2/1/1; C3: 1/2/1, C4:1/1/2 was evaluated microdilution assay. The various interactions of OEs in the different combinations were determined then the OE and the most active combination were retained to determine their kinetic parameters on S. flexneri. Results: Antibiotic susceptibility tests revealed that most Shigella isolates (n = 4) were resistant to six antibiotics tested. Ciprofloxacin (40%), Nalidixic acid (60%), Tetracycline (80%), Amoxicillin (100%), Cefotaxime (80%), Erythromycin (100%), and Cotrimoxazole (80%) were the profiles found in the different strains of Shigella. About the antibacterial activity of OEs, Drypetes gossweileriOE and C2 combination had shown a higher Shigellicide property with a Minimal Inhibitory Concentration(MIC) respectivelyranging from 0.078 mg/mL to 0.312 mg/mL and 0.012 to 1.562 mg/mL. Combinations of OEs showed various interactions whose synergistic effects were mostly encountered. The best deactivation was obtained by the combination C2 at 16 MIC withb= 1.962. Conclusion: the susceptibility of Shigella to OEs and their combinations justifies their use in traditional medicine in the treatment of shigellosis.

Keywords: shigella, multidrug-resistant, EOs, kinetic

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Authors: Suelen C. Lima, André C. de Oliveira, Rosemary A. Roque

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Dengue is fatal arboviruses transmitted by the A. aegypti mosquito to more than 100 countries, for which the WHO estimates that 2.5 million people will be infected by these disease. The widespread of these diseases is due, among other factors, to the resistance that A. aegypti has to several commercial insecticides. On the other hand, natural products based on plants of the genus Piper (Piperaceae) are characterized by their insecticidal activities against mosquitoes. Piper longepetiolatum and Piper brachypetiolatum are species with wide distribution in the State of Amazonas. However, there is no investigation of phytochemical or biological of these plants against mosquitoes such as A. aegypti. The main of this study was to identify the chemical composition of the essential oil (EOs) from P. longepetiolatum and P. brachypetiolatum and to evaluate the biological activity against A. aegypti. The EOs were extracted by hydrodistillation from leaves (200 g) of P. longepetiolatum and P. brachypetiolatum and analyzed by GC-MS and GC-FID. The main compounds β-caryophyllene (99.9% of purity) and E-nerolidol (99.4% of purity) were purchased from Sigma-Aldrich® Brazil. The larvicidal activity of EOs (20 to 100 ppm), β-caryophyllene and E-nerolidol (10 to 50 ppm) was performed according to WHO protocol against A. aegypti larvae. The GC-MS and GC-FID analysis of EOs from P. longepetiolatum and P. brachypetiolatum indicated the majority presence of β-caryophyllene (35.42%) and E-nerolidol (49.79%), respectively. The results showed that all natural products presented larvicidal activity against A. aegypti. In this aspect, the OE from P. brachypetiolatum (LC50 of 15.51 ppm and LC90 of 22.79 ppm) was more active than the OE from P. longepetiolatum (LC50 of 47.17 ppm and LC90 of 69.60 ppm) (p < 0.05). Regarding of main compounds, E-nerolidol (LC50 of 9.50 ppm and LC90 of 23.89 ppm) showed higher larvicidal activity than the β-caryophyllene compound (LC50 of 79.00 ppm and LC90 of 230.91 ppm) (p < 0.05). The larvae treated with these natural products showed tremors and lethargic movements, suggesting that these natural products have neurotoxic action. These observations support studies to investigate the mechanism of action. This is the first record of the chemical composition and larvicidal activity of the EO from P. longepetiolatum and P. brachypetiolatum rich in β-caryophyllene and E-nerolidol against A. aegypti larvae.

Keywords: piperaceae, aedes, sesquiterpenes, biological control

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Authors: Zhala Ahmad, Zainab Lazim, Haider Hamzah

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Bacterial resistance is a pressing global health issue, with multidrug-resistant (MDR), extensively drug-resistant (XDR), and pandrug-resistant (PDR) strains to pose a serious threat. In this context, researchers are investigating effective, safe, and affordable metabolites to combat these pathogens. This study focuses on gum essential oil (GEO) extracted from Pistacia atlantica and its activity and the mechanism of action against XDR Gram-negative Acinetobacter baumannii. GEO was extracted by hydrodistillation and analyzed using GC-MS. Eleven A. baumannii isolates were collected from the ward environment of Burn and Plastic Surgery Hospital in Al Sulaymaniyah City, Iraq. They were identified using the VITEK 2 system, 16S rRNA gene, and confirmed with the blaₒₓₐ₋₅₁ gene; A. baumannii ATCC 19606 was used as a reference strain. The isolates were identified as resistant to twelve different antibiotics spanning six distinct antibiotic classes while showing susceptibility to tetracycline and trimethoprim. Over 40 chemical constituents were detected in the gum's essential oils, with α-pinene being the most abundant. GEO was found to inhibit the growth of A. baumannii isolates; the minimum inhibitory concentration (MIC) of GEO was 2.5 µl/ml. GEO induced protein leakage, phosphate, and potassium ion efflux, distorted cell morphology, and cell death in the tested bacteria. GEO exhibited bacterial clearance and anti-adhesion activity using Band-Aids. This study's findings suggest that GEO could be used as a potential alternative treatment for infectious diseases caused by XRD pathogens, shedding further light on the importance of GEO in biomedical applications. Future studies must focus on generating clinically feasible sources of GEO for testing in small animal models before proceeding to human trials, ensuring safe and effective translation from the laboratory to the clinic.

Keywords: antibiotic resistance, Acinetobacter baumannii, essential oils, Pistacia atlantica, alpha-pinene

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Authors: A.Assariotakis, P. Vahamidis, P. Tarantilis, G. Economou

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Greece, due to its geographical location and the particular climatic conditions, presents high biodiversity of Medicinal and Aromatic Plants. Among them, the genus Origanum not only presents a wide distribution, but it also has great economic importance. After extensive surveys in Ikaria Island (Greece), 3 species of the genus Origanum were identified, namely, Origanum vulgare ssp. hirtum (Greek oregano), Origanum onites (Turkish oregano) and Origanum x intercedens (hybrid), a naturally occurring hybrid between O. hirtum and O. onites. The purpose of this study was to determine their morphological as well as their temporal variability in essential oil yield and composition under field conditions. For this reason, a plantation of each species was created using vegetative propagation and was established at the experimental field of the Agricultural University of Athens (A.U.A.). From the establishment year and for the following two years (3 years of observations), several observations were taken during each growing season with the purpose of identifying the morphological differences among the studied species. Each year collected plant (at bloom stage) material was air-dried at room temperature in the shade. The essential oil content was determined by hydrodistillation using a Clevenger-type apparatus. The chemical composition of essential oils was investigated by Gas Chromatography-Mass Spectrometry (GC – MS). Significant differences were observed among the three oregano species in terms of plant height, leaf size, inflorescence features, as well as concerning their biological cycle. O. intercedens inflorescence presented more similarities with O. hirtum than with O. onites. It was found that calyx morphology could serve as a clear distinction feature between O. intercedens and O. hirtum. The calyx in O. hirtum presents five isometric teeth whereas in O. intercedens two high and three shorter. Essential oil content was significantly affected by genotype and year. O. hirtum presented higher essential oil content than the other two species during the first year of cultivation, however during the second year the hybrid (O. intercedens) recorded the highest values. Carvacrol, p-cymene and γ-terpinene were the main essential oil constituents of the three studied species. In O. hirtum carvacrol content varied from 84,28 - 93,35%, in O. onites from 86,97 - 91,89%, whereas in O. intercedens it was recorded the highest carvacrol content, namely from 89,25 - 97,23%.

Keywords: variability, oregano biotypes, essential oil, carvacrol

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Authors: Randa M. T. Mohamed

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Spices have long been used as traditional ingredients in the kitchen for seasoning, coloring, aromatic and food preservative properties. Besides, spices are equally used for therapeutic purposes. The objective of this study was to survey and document the medicinal properties of spices commonly used in the Sudanese kitchen for different food preparations. Also, extracts from reported spices were screened for the presence of secondary metabolites as well as their antioxidant and beta-lactamase inhibitory properties. This study was conducted in the Rekabbya Quartier in Omdurman, Khartoum State, Sudan. Information was collected by carrying out semi-structured interviews. All informants (30) in the present study were women. Spices were purchased from Attareen shop in Omdurman. Essential oils from spices were extracted by hydrodistillation and ethanolic extracts by maceration. Phytochemical screening was performed by thin layer chromatography (TLC). The antioxidant capacity of essential oils and ethanolic extracts was investigated through TLC bioautography. Beta lactamase inhibitory activity was performed by the acidimetric test. Ethnobotany study showed that a total of 16 spices were found to treat 36 ailments belonging to 10 categories. The most frequently claimed medicinal uses were for the digestive system diseases treated by 14 spices and respiratory system diseases treated by 8 spices. Gynaecological problems were treated by 4 spices. Dermatological diseases were cured by 5 spices while infections caused by tapeworms and other microbes causing dysentery were treated by 3 spices. 4 spices were used to treat bad breath, bleeding gum and toothache. Headache, eyes infection, cardiac stimulation and epilepsy were treated by one spice each. Other health problem like fatigue and loss of appetite and low breast milk production were treated by 1, 3 and 2 spices respectively. The majority (69%, 11/16) of spices were exported from different countries like India, China, Indonesia, Ethiopia, Egypt and Nigeria while 31% (5/16) was cultivated in Sudan. Essential oils of all spices were rich in terpenes while ethanolic extracts contained variable classes of secondary metabolites. Both essential oils and ethanolic extracts of all spices exerted considerable antioxidant activity. Only one extract, Syzygium aromaticum, possessed beta lactamase inhibitory activity. In conclusion, this study could contribute in conserving information on traditional medicinal uses of spices in Sudan. Also, the results demonstrated the potential of some of these spices to exert beneficial antimicrobial and antioxidant effect. Detailed phytochemical and biological assays of these spices are recommended.

Keywords: spices, ethnobotany, phytoconstituents, antioxidant, beta lactamase inhibition

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Authors: Igor Jerkovic

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Biodiversity of flora provides many different nectar sources for the bees. Unifloral honeys possess distinctive flavours, mainly derived from their nectar sources (characteristic volatile organic components (VOCs)). Specific or nonspecific VOCs (chemical markers) could be used for unifloral honey characterisation as addition to the melissopalynologycal analysis. The main honey volatiles belong, in general, to three principal categories: terpenes, norisoprenoids, and benzene derivatives. Some of these substances have been described as characteristics of the floral source, and other compounds, like several alcohols, branched aldehydes, and furan derivatives, may be related to the microbial purity of honey processing and storage conditions. Selection of the extraction method for the honey volatiles profiling should consider that heating of the honey produce different artefacts and therefore conventional methods of VOCs isolation (such as hydrodistillation) cannot be applied for the honey. Two-way approach for the isolation of the honey VOCs was applied using headspace solid-phase microextraction (HS-SPME) and ultrasonic solvent extraction (USE). The extracts were analysed by gas chromatography and mass spectrometry (GC-MS). HS-SPME (with the fibers of different polarity such as polydimethylsiloxane/ divinylbenzene (PDMS/DVB) or divinylbenzene/carboxene/ polydimethylsiloxane (DVB/CAR/PDMS)) enabled isolation of high volatile headspace VOCs of the honey samples. Among them, some characteristic or specific compounds can be found such as 3,4-dihydro-3-oxoedulan (in Centaurea cyanus L. honey) or 1H-indole, methyl anthranilate, and cis-jasmone (in Citrus unshiu Marc. honey). USE with different solvents (mainly dichloromethane or the mixture pentane : diethyl ether 1 : 2 v/v) enabled isolation of less volatile and semi-volatile VOCs of the honey samples. Characteristic compounds from C. unshiu honey extracts were caffeine, 1H-indole, 1,3-dihydro-2H-indol-2-one, methyl anthranilate, and phenylacetonitrile. Sometimes, the selection of solvent sequence was useful for more complete profiling such as sequence I: pentane → diethyl ether or sequence II: pentane → pentane/diethyl ether (1:2, v/v) → dichloromethane). The extracts with diethyl ether contained hydroquinone and 4-hydroxybenzoic acid as the major compounds, while (E)-4-(r-1’,t-2’,c-4’-trihydroxy-2’,6’,6’-trimethylcyclo-hexyl)but-3-en-2-one predominated in dichloromethane extracts of Allium ursinum L. honey. With this two-way approach, it was possible to obtain a more detailed insight into the honey volatile and semi-volatile compounds and to minimize the risks of compound discrimination due to their partial extraction that is of significant importance for the complete honey profiling and identification of the chemical biomarkers that can complement the pollen analysis.

Keywords: honey chemical biomarkers, honey volatile compounds profiling, headspace solid-phase microextraction (HS-SPME), ultrasonic solvent extraction (USE)

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Authors: Maged El-Sayed Mohamed

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Eucalyptus camaldulensis is one on the most well-known species of the genus Eucalyptus in the Middle east, its importance relay on the high production of its unique volatile constituents which exhibits many medicinal and pharmacological activities. The gall-forming wasp (Leptocybe invasa) has recently come into sight as the main pest attacking E. camaldulensis and causing severe injury. The wasp lays its eggs in the petiole and midrib of leaves and stems of young shoots of E. camaldulensis, which leads to gall formation. Gall formation by L. invasa damages growing shoot and leaves of Eucalyptus, resulting in abscission of leaves and drying. AIM: This study is an attempt to investigate the effect of the gall wasp (Leptocybe invasa) attack on the volatile constitutes of E. camaldulensis. This could help in the control of this wasp through stimulating plant defenses or production of a new allelochemicals or insecticide. The study of volatile constitutes of Eucalyptus before and after attack by the wasp can help the re-use and recycle of the infected Eucalyptus trees for new pharmacological and medicinal activities. Methodology: The fresh gall wasp-attacked and healthy leaves (100 g each) were cut and immediately subjected to hydrodistillation using Clevenger-type apparatus for 3 hours. The volatile fractions isolated were analyzed using Gas chromatography/mass spectrometry (GC/MS). Kovat’s retention indices (RI) were calculated with respect to a set of co-injected standard hydrocarbons (C10-C28). Compounds were identified by comparing their spectral data and retention indices with Wiley Registry of Mass Spectral Data 10th edition (April 2013), NIST 11 Mass Spectral Library (NIST11/2011/EPA/NIH) and literature data. Results: Fifty-nine components representing 89.13 and 88.60% of the total volatile fraction content respectively were quantitatively analyzed. Twenty-six major compounds at an average concentration greater than 0.1 ± 0.02% have been used for the statistical comparison. From those major components, twenty-one were found in both the attacked and healthy Eucalyptus leaves’ fractions in different concentration and five components, mono terpene p-Mentha-2-4(8) diene and the sesquiterpenes δ-elemene, β-elemene, E-caryophyllene and Bicyclogermacrene, were unique and only produced in the attacked-leaves’ fraction. CONCLUSION: Newly produced components or those commonly found in the volatile fraction and changed in concentration could represent a part of the plant defense mechanisms or might be an element of the plant allelopathic and communication mechanisms. Identification of the components of the gall wasp-damaged leaves can help in their recycling for different physiological, pharmacological and medicinal uses.

Keywords: Eucalyptus camaldulensis, eucalyptus recycling, gall wasp, Leptocybe invasa, plant defense mechanisms, Terpene fraction

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Authors: Fatima Bouazza, Khadija Khedid, Lamiae Amallah, Aziz Mouhaddach, Basma Boukour, Jihane Ennadir, Rachida Hassikou

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In order to combine milk and its derived products conservation and flavoring, Moroccans often used aromatic and medicinal plants. These plant extracts are endowed with several nutritive and therapeutic properties. This study constitutes a first national assessment of physico-chemical quality of sheep’s milk from moroccan Sardi breed and the evaluation of the antibacterial effect of three medicinal plants extracts: Aloe barbadensis Miller, Thymus satureioides and Mentha pulegium on flora isolated from this sheep's milk. 100 milk samples were collected in four regions of Morocco. The bacteria isolated were identified by classical and molecular methods (16S rRNA sequencing) and tested, according to the disk method, for their sensitivity to several antibiotics. The physico-chemical analyzes of sheep’s milk concerned the pH, titratable acidity, density, dry extract, freezing point and contents of: fat, proteins, lactose and calcium. The essential oils (EOs) of T. satureioides and M .pulegium were extracted by hydrodistillation and analyzed by GC / MS, while the Aloe vera leaf pulp was analyzed by the methods of Harborne and HPLC. A total number of 125 bacteria have been identified. Significant resistance to chemical antibiotics has been noted in LABs. The average temperature value of milk is around 57.15 °C, the pH is 6.56, the titratable acidity is around 3.4 ° D, the density is 1.035g / cm³ , the total dry extract is around 169.5g / l, the ash (9.8g / l), the freezing point (- 0.556 °C) while the average fat content is 67.85g / l . The samples richest in fat belong to the region of Settat, cradle of the Sardi breed, with a maximum average value of 74.4g / l. The average protein is 56g / l, lactose (39.92g / l), and calcium (1.855g / l). Analysis of the major components of EOs revealed the dominance of borneol in the case of T. satureioides and of pulegone in M. pulegium. Aloe vera gel contains alkaloids, flavonoids, catechic tannins, saponins and 1.60 µg / ml of aloin. The plant extracts have a bactericidal effect on E. coli, Klebsiellaoxytoca and Staphylococci and bacteriostatic effect on LABs of technological interest (Lactobacillus). As a result of this study, it is believed that the consumption of sardi sheep’s milk would be of nutritional benefit. Its richness in fat and proteins predisposes it for biotechnological development in the manufacture of cheese and yogurt. Also, the use of aromatic and medicinal plants, as natural additives would be of great benefit to flavor and maintain its quality.

Keywords: sheep’s milk, lactic flora, antimicrobial power, aloe barbadensis miller, thymus satureioides, mentha pulegium

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Authors: Victor Pena Ribeiro, Caroline Arruda, Jennyfer Andrea Aldana Mejia, Jairo Kenupp Bastos

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Leishmaniasis is a serious health problem around the world. The treatment of infected individuals with pentavalent antimonial drugs is the main therapeutic strategy. However, they present high toxicity and persistence side effects. Therefore, the discovery of new and safe natural-derived therapeutic agents against leishmaniasis is important. Propolis is a resin of viscous consistency produced by Apis mellifera bees from parts of plants. The main types of Brazilian propolis are green, red, yellow and brown. Thus, the aim of this work was to investigate the chemical composition and leishmanicidal properties of a brown propolis (BP). For this purpose, the hydroalcoholic crude extract of BP was obtained and was fractionated by liquid-liquid chromatography. The chemical profile of the extract and its fractions were obtained by HPLC-UV-DAD. The fractions were submitted to preparative HPLC chromatography for isolation of the major compounds of each fraction. They were analyzed by NMR for structural determination. The volatile compounds were obtained by hydrodistillation and identified by GC/MS. Promastigote forms of Leishmania amazonensis were cultivated in M199 medium and then 2×106 parasites.mL-1 were incubated in 96-well microtiter plates with the samples. The BP was dissolved in dimethyl sulfoxide (DMSO) and diluted into the medium, to give final concentrations of 1.56, 3.12, 6.25, 12.5, 25 and 50 µg.mL⁻¹. The plates were incubated at 25ºC for 24 h, and the lysis percentage was determined by using a Neubauer chamber. The bioassays were performed in triplicate, using a medium with 0.5% DMSO as a negative control and amphotericin B as a positive control. The leishimnicidal effect against promastigote forms was also evaluated at the same concentrations. Cytotoxicity experiments also were performed in 96-well plates against normal (CHO-k1) and tumor cell lines (AGP01 and HeLa) using XTT colorimetric method. Phenolic compounds, flavonoids, and terpenoids were identified in brown propolis. The major compounds were identified as follows: p-coumaric acid (24.6%) for a methanolic fraction, Artepelin-C (29.2%) for ethyl acetate fraction and the compounds of hexane fraction are in the process of structural elucidation. The major volatile compounds identified were β-caryophyllene (10.9%), germacrene D (9.7%), nerolidol (10.8%) and spathulenol (8.5%). The propolis did not show cytotoxicity against normal cell lines (CHO) with IC₅₀ > 100 μg.mL⁻¹, whereas the IC₅₀ < 10 μg.mL⁻¹ showed a potential against the AGP01 cell line, propolis did not demonstrate cytotoxicity against HeLa cell lines IC₅₀ > 100 μg.mL⁻¹. In the determination of the leishmanicidal activity, the highest (50 μg.mL⁻¹) and lowest (1.56 μg.mL⁻¹) concentrations of the crude extract caused the lysis of 76% and 45% of promastigote forms of L. amazonensis, respectively. To the amastigote form, the highest (50 μg.mL⁻¹) and lowest (1.56 μg.mL⁻¹) concentrations caused the mortality of 89% and 75% of L. amazonensis, respectively. The IC₅₀ was 2.8 μg.mL⁻¹ to amastigote form and 3.9 μg.mL⁻¹ to promastigote form, showing a promising activity against Leishmania amazonensis.

Keywords: amastigote, brown propolis, cytotoxicity, promastigote

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Authors: Dmitriy Yu. Korulkin, Raissa A. Muzychkina

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The last decade the growth of severe and disseminated forms of inflammatory diseases is observed in Kazakhstan, in particular, septic shock, which progresses on 3-15% of patients with infectious complications of postnatal period. In terms of the rate of occurrence septic shock takes third place after hemorrhagic and cardiovascular shock, in terms of lethality it takes first place. The structure of obstetric sepsis has significantly changed. Currently the first place is taken by postabortive sepsis (40%) that is connected with usage of imperfect methods of artificial termination of pregnancy in late periods (intraamnial injection of sodium chloride, glucose). The second place is taken by postnatal sepsis (32%); the last place is taken by septic complications of caesarean section (28%). In this connection, search for and assessment of effectiveness of new medicines for treatment of postoperative infectious complications, having biostimulating effect and speeding up regeneration processes, is very promising and topical. Essential oil was obtained by the method hydrodistillation air-dry aerial part of Sedum L. plants using Clevenger apparatus. Pilot batch of plant medicinal product based on Sedum essential oils was produced by Chimpharm JSC, Santo Member of Polpharma Group (Kazakhstan). During clinical test of the plant medicinal product based on Sedum L. essential oils 37 female patients at the age from 35 to 57 with clinical signs of complicated postoperative processes and 12 new mothers with clinical signs of inflammatory process on sutures on anterior abdominal wall after caesarean section and partial disruption of surgical suture line on perineum were examined. Medicine usage methods - surgical wound treatment 2 times a day, treatment with other medicines of local action was not performed. Before and after treatment general clinical test, determination of immune status, bacterioscopic test of wound fluid was performed to all women, medical history data was taken into account, wound cleansing and healing time, full granulations, side effects and complications, satisfaction with the used medicine was assessed. On female patients with inflammatory infiltration and partial disruption of surgical suture line anesthetic wound healing effect of plant medicinal product based on Sedum L. essential oils was observed as early as on the second day after beginning of using it, wound cleansing took place, as a rule, within the first row days. Hyperemia in the area of suture line also was not observed for 2-3-d day of usage of medicine, good constant course was observed. The absence of clinical effect on this group of patients was not registered. The represented data give evidence of that clinical effect was accompanied with normalization of changed laboratory findings. No allergic responses or side effects were observed during usage of the plant medicinal products based on Sedum L. essential oils.

Keywords: antiinflammatory, bioactive substances, essential oils, isolation, sedum L., wound healing

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Authors: Dmitry Yu. Korulkin, Raissa A. Muzychkina

Abstract:

Bioactive substances of plant origin can be one of the advanced means of solution to the issue of combined therapy to inflammation. The main advantages of medical plants are softness and width of their therapeutic effect on an organism, the absence of side effects and complications even if the used continuously, high tolerability by patients. Moreover, medial plants are often the only and (or) cost-effective sources of natural biologically active substances and medicines. Along with other biologically active groups of chemical compounds, essential oils with wide range of pharmacological effects became very ingrained in medical practice. Essential oil was obtained by the method hydrodistillation air-dry aerial part of Polygonum L. plants using Clevenger apparatus. Qualitative composition of essential oils was analyzed by chromatography-mass-spectrometry method using Agilent 6890N apparatus. The qualitative analysis is based on the comparison of retention time and full mass-spectra with respective data on components of reference oils and pure compounds, if there were any, and with the data of libraries of mass-spectra Wiley 7th edition and NIST 02. The main components of essential oil are for: Polygonum amphibium L. - γ-terpinene, borneol, piperitol, 1,8-cyneole, α-pinene, linalool, terpinolene and sabinene; Polygonum minus Huds. Fl. Angl. – linalool, terpinolene, camphene, borneol, 1,8-cyneole, α-pinene, 4-terpineol and 1-octen-3-ol; Polygonum alpinum All. – camphene, sabinene, 1-octen-3-ol, 4-carene, p- and o-cymol, γ-terpinene, borneol, -terpineol; Polygonum persicaria L. - α-pinene, sabinene, -terpinene, 4-carene, 1,8-cyneole, borneol, 4-terpineol. Antibacterial activity was researched relating to strains of gram-positive bacteria Staphylococcus aureus, Bacillus subtilis, Streptococcus agalacticae, relating to gram-negative strain Escherichia coli and to yeast fungus Сandida albicans using agar diffusion method. The medicines of comparison were gentamicin for bacteria and nystatin for yeast fungus Сandida albicans. It has been shown that Polygonum L. essential oils has moderate antibacterial effect to gram-positive microorganisms and weak antifungal activity to Candida albicans yeast fungus. At the second stage of our researches wound healing properties of ointment form of 3% essential oil was researched on the model of flat dermal wounds. To assess the influence of essential oil on healing processes the model of flat dermal wound. The speed of wound healing on rats of different groups was judged based on assessment the area of a wound from time to time. During research of wound healing properties disturbance of integral in neither group: general condition and behavior of animals, food intake, and excretion. Wound healing action of 3% ointment on base of Polygonum L. essential oil and polyethyleneglycol is comparable with the action of reference substances. As more favorable healing dynamics was observed in the experimental group than in control group, the tested ointment can be deemed more promising for further detailed study as wound healing means.

Keywords: antibacterial, antifungal, bioactive substances, essential oils, isolation, Polygonum L.

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Authors: S. Mamoucha, N.Tsafantakis, T. Ioannidis, S. Chatzipanagiotou, C. Nikolaou, L. Skaltsounis, N. Fokialakis, N. Christodoulakis

Abstract:

Introduction: Pistacia lentiscus L. (well known as Mastic tree) is an evergreen sclerophyllous shrub that extensively thrives in the eastern Mediterranean area yet only the trees cultivated in the southern region of the Greek island Chios produces mastic resin. Different parts of P. lentiscus L. var. chia have been used in folk medicine for various purposes, such as tonic, aphrodisiac, antiseptic, antihypertensive and management of dental, gastrointestinal, liver, urinary, and respiratory tract disorders. Several studies have focused on the antibacterial activity of its resin (gum) and its essential oil. However, there is no study combining anatomy of the plant organs, phytochemical profile, and antibacterial screening of the plant. In our attempt to discover novel bioactive metabolites from the mastic tree, we screened its antibacterial activity not only against ATCC strains but also against clinical, resistant strains. Materials-methods: Leaves were investigated using Transmission (ΤΕΜ) and Scanning Εlectron Microscopy (SEM). Histochemical tests were performed on fresh and fixed tissue. Extracts prepared from dried, powdered leaves using 3 different solvents (DCM, MeOH and H2O) the waste water obtained after a hydrodistillation process for essential oil production were screened for their phytochemical content and antibacterial activity. Μetabolite profiling of polar and non-polar extracts was recorded by GC-MS and LC-HRMS techniques and analyzed using in-house and commercial libraries. The antibacterial screening was performed against Staphylococcus aureus ATCC25923, Escherichia coli ATCC25922, Pseudomonas aeruginosa ATCC27853 and against clinical, resistant strains Methicillin-resistant S. aureus (MRSA), Carbapenem-Resistant Metallo-β-Lactamase (carbapenemase) P. aeruginosa (VIM), Klebsiella pneumoniae carbapenemases (KPCs) and Acinetobacter baumanii resistant strains. The antibacterial activity was tested by the Kirby Bauer and the Agar Well Diffusion method. The zone of inhibition (ZI) of each extract was measured and compared with those of common antibiotics. Results: Leaf is compact with inosclereids and numerous idioblasts containing a globular, spiny crystal. The major nerves of the leaf contain a resin duct. Mesophyll cells showed accumulation of osmiophillic metabolites. Histochemical treatments defined secondary metabolites in subcellular localization. The phytochemical investigation revealed the presence of a large number of secondary metabolites, belonging to different chemical groups, such as terpenoids, phenolic compounds (mainly myricetin, kaempferol and quercetin glycosides), phenolic, and fatty acids. Among the extracts, the hydrostillation wastewater achieved the best results against most of the bacteria tested. MRSA, VIM and A. baumanii were inhibited. Conclusion: Extracts from plants have recently been of great interest with respect to their antimicrobial activity. Their use emerged from a growing tendency to replace synthetic antimicrobial agents with natural ones. Leaves of P. lentiscus L. var. chia showed a high antimicrobial activity even against drug - resistant bacteria. Future prospects concern the better understanding of mode of action of the antibacterial activity, the isolation of the most bioactive constituents and the clarification if the activity is related to a single compound or to the synergistic effect of several ones.

Keywords: antibacterial screening, leaf anatomy, phytochemical profile, Pistacia lentiscus var. chia

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