Chemical and biological characterization of Melaleuca subulata (Cheel) Craven leaves' volatile constituents supported by chemometric analysis and molecular docking.

BACKGROUND: The genus Melaleuca (Myrtaceae) comprises dozens of essential oil (EO)-rich species that are appreciated worldwide for their various medicinal values. Additionally, they are renowned in traditional medicine for their antimicrobial, antifungal, and other skin-related activities. The current study investigated the chemical profile and skin-related activities of volatile constituents derived from M. subulata (Cheel) Craven (Synonym Callistemon subulatus) leaves cultivated in Egypt for the first time. METHODS: The volatile components were extracted using hydrodistillation (HD), headspace (HS), and supercritical fluid (SF). GC/MS and Kovat's retention indices were implemented to identify the volatile compounds, while the variations among the components were assessed using Principal Component Analysis and Hierarchical Cluster Analysis. The radical scavenging activity was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), oxygen radical absorbance capacity (ORAC) and ß-carotene assays. Moreover, the anti-aging effect was evaluated using anti-elastase, and anti-collagenase, while the antimicrobial potential was deduced from the agar diffusion and broth microdilution assays. Lastly, the molecular docking study was executed using C-docker protocol in Discovery Studio 4.5 to rationalize the binding affinity with targeted enzymes. RESULTS: The SF extraction approach offered the highest EO yield, being 0.75%. According to the GC/MS analysis, monoterpene hydrocarbons were the most abundant volatile class in the HD oil sample (54.95%), with α-pinene being the most copious component (35.17%). On the contrary, the HS and SF volatile constituents were pioneered with oxygenated monoterpenes (72.01 and 36.41%) with eucalyptol and isopulegone being the most recognized components, representing 67.75 and 23.46%, respectively. The chemometric analysis showed segregate clustering of the three extraction methods with α-pinene, eucalyptol, and isopulegone serving as the main discriminating phytomarkers. Concerning the bioactivity context, both SF and HD-EOs exhibited antioxidant effects in terms of ORAC and ß-carotene bleaching. The HD-EO displayed potent anti-tyrosinase activity, whereas the SF-EO exhibited significant anti-elastase properties. Moreover, SF-EO shows selective activity against gram-positive skin pathogens, especially S. aureus. Ultimately, molecular docking revealed binding scores for the volatile constituents; analogous to those of the docked reference drugs. CONCLUSIONS: M. subulata leaves constitute bioactive volatile components that may be indorsed as bioactive hits for managing skin aging and infection, though further in vivo studies are recommended.

Inhaled Litsea glaucescens K. (Lauraceae) leaves' essential oil has anxiolytic and antidepressant-like activity in mice by BDNF pathway activation.

ETHNOPHARMACOLOGICAL RELEVANCE: Litsea glaucescens K. (Lauraceae) is a small tree from the Mexican and Central American temperate forests, named as "Laurel". Its aromatic leaves are ordinarily consumed as condiments, but also are important in Mexican Traditional Medicine, and among the most important non wood forest products in this area. The leaves are currently used in a decoction for the relief of sadness by the Mazahua ethnic group. Interestingly, "Laurel" has a long history. It was named as "Ehecapahtli" (wind medicine) in pre-Columbian times and applied to heal maladies correlated to the Central Nervous System, among them depression, according to botanical texts written in the American Continent almost five centuries ago. AIM OF THE STUDY: Depression is the first cause of incapacity in the world, and society demands alternative treatments, including aromatherapy. We have previously demonstrated the antidepressant-like activity of L. glaucescens leaves' essential oil (LEO), as well as their monoterpenes linalool, and beta-pinene by intraperitoneal route in a mice behavioral model. Here we now examined if LEO and linalool exhibit this property and anxiolytic activity when administered to mice by inhalation. We also investigated if these effects occur by BDNF pathway activation in the brain. MATERIALS AND METHODS: The LEO was prepared by distillation with water steam and analyzed by gas chromatography-mass spectrometry (GC-MS). The monoterpenes linalool, eucalyptol and ß-pinene were identified and quantified. Antidepressant type properties were determined with the Forced Swim Test (FST) on mice previously exposed to LEO or linalool in an inhalation chamber. The spontaneous locomotor activity and the sedative effect were assessed with the Open Field Test (OFT), and the Exploratory Cylinder (EC), respectively. The anxiolytic properties were investigated with the Elevated Plus Maze Apparatus (EPM) and the Hole Board Test (HBT). All experiments were video documented. The mice were subjected to euthanasia, and the brain hippocampus and prefrontal cortex were dissected. RESULTS: The L. glaucescens essential oil (LEO) contains 31 compounds according to GC/MS, including eucalyptol, linalool and beta-pinene. The LEO has anxiolytic effect by inhalation in mice, as well as linalool, and ß-pinene, as indicated by OFT and EC tests. The LEO and imipramine have antidepressant like activity in mice as revealed by the FST; however, linalool and ketamine treatments didn't modify the time of immobility. The BDNF was increased in FST in mice treated with LEO in both areas of the brain as revealed by Western blot; but did not decrease the level of corticosterone in plasma. The OFT indicated that LEO and imipramine didn't reduce the spontaneous motor activity, while linalool and ketamine caused a significant decrease. CONCLUSION: Here we report by the first time that L. glaucescens leaves essential oil has anxiolytic effect by inhalation in mice, as well as linalool, and ß-pinene. This oil also maintains its antidepressant-like activity by this administration way, similarly to the previously determined intraperitoneally. Since inhalation is a common administration route for humans, our results suggest L. glaucescens essential oil deserve future investigation due to its potential application in aromatherapy.

Attenuation of quorum sensing regulated virulence functions and biofilm of pathogenic bacteria by medicinal plant Artemisia annua and its phytoconstituent 1, 8-cineole.

The emergence of multidrug resistance (MDR) in bacterial pathogens is a serious public health concern. A significant therapeutic target for MDR infections is the quorum sensing-regulated bacterial pathogenicity. Determining the anti-quorum sensing abilities of certain medicinal plants against bacterial pathogens as well as the in-silico interactions of particular bioactive phytocompounds with QS and biofilm-associated proteins were the objectives of the present study. In this study, 6 medicinal plants were selected based on their ethnopharmacological usage, screened for Anti-QS activity and Artemisia annua leaf extract (AALE) demonstrated pigment inhibitory activity against Chromobacterium violaceum CV12472. Further, the methanol active fraction significantly inhibited the virulence factors (pyocyanin, pyoverdine, rhamnolipid and swarming motility) of Pseudomonas aeruginosa PAO1 and Serratia marcescens MTCC 97 at respective sub-MICs. The inhibition of biofilm was determined using a microtiter plate test and scanning electron microscopy. Biofilm formation was impaired by 70%, 72% and 74% in P. aeruginosa, C. violaceum and S. marcescens, respectively at 0.5xMIC of the extract. The phytochemical content of the extract was studied using GC-MS and 1, 8-cineole was identified as major bioactive compound. Furthermore, 1, 8-cineole was docked with quorum sensing (QS) proteins (LasI, LasR, CviR, and rhlR) and biofilm proteins (PilY1 and PilT). In silico docking and dynamics simulations studies suggested interactions with QS-receptors CviR', LasI, LasR, and biofilm proteins PilY1, PilT for anti-QS activity. Further, 1, 8-cineole demonstrated 66% and 51% reduction in violacein production and biofilm formation, respectively to validate the findings of computational analysis. Findings of the present investigation suggests that 1, 8-cineole plays a crucial role in the QS and biofilm inhibitory activity demonstrated by Artemisia annua extract. RESEARCH HIGHLIGHTS: Artemisia annua leaf extract (AALE) methanol fraction demonstrated broad-spectrum QS and biofilm inhibition Scanning electron microscopy (SEM) confirmed biofilm inhibition Molecular docking and simulation studies suggested positive interactions of 1,8-cineol with QS-receptors and biofilm proteins.

Phytochemical investigation of Pistacia lentiscus L. var. Chia leaves: A byproduct with antimicrobial potential.

Pistacia lentiscus L. var. Chia belongs to the Anacardiaceae family, and it is cultivated only in the south part of Chios island, in Greece. Even though it is renowned for its unique resin, Chios mastic gum (CMG), the tree leaves have also been used in traditional medicine, while the annual pruning generates a large biomass of unused by-products. Thus, the aim of the present study was the detailed phytochemical investigation of P. lentiscus var. Chia leaves towards the search of antimicrobial agents. UPLC-HRMS & HRMS/MS based dereplication methods led to the detailed characterization of the aqueous leaf extract. In addition, twelve compounds were isolated and purified from the methanol extract and were identified using spectroscopic and spectrometric methods (NMR, HRMS) belonging to phenolic acids, tannins, flavonoids and terpenes, with the most interesting being 2-hydroxy-1,8-cineole ß-D-glucopyranoside which was isolated for the first time in the Anacardiaceae family. Remarkably, based on NMR data, methanol and aqueous extracts were found to be particularly rich in shikimic acid, a valuable building block for the pharmaceutical industry, for instance in the synthesis of the active ingredient of Tamiflu®, oseltamivir. Finally, extracts (EtOAc, MeOH, H2O) and major compounds i.e., shikimic acid, 2-hydroxy-1,8-cineole ß-D-glucopyranoside and myricitrin were evaluated for their antimicrobial properties. MeOH and H2O mastic leaf extracts as well as myricitrin and, particularly, 2-hydroxy-1,8-cineole ß-D-glucopyranoside showed significant selective activity against pathogenic Mucorales, but not against Aspergilli (Aspergillus nidulans, Aspergillus fumigatus), Candida albicans or bacteria (Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis).

Cineole inhibits the biosynthesis of leukotrienes and prostaglandins to alleviate allergic rhinitis: Insights from metabolomics.

Allergic rhinitis (AR) is a common allergic disease characterized by nasal congestion, rhinorrhoea, and sneezing. Cineole, a monoterpenoid compound widely present in various volatile oils, has a wide range of pharmacological activities and is of interest in allergic airway diseases for its anti-inflammatory and anti-mucus production abilities. However, the protective effects of cineole in mice with allergic rhinitis and its mechanisms have not been well investigated. In this study, the protective effect of cineole against ovalbumin-induced (OVA-induced) allergic rhinitis and its molecular mechanism is investigated by metabolomic analysis based on ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). OVA combined with aluminum hydroxide adjuvant is used to sensitize and establish the allergic rhinitis (AR) mouse model. The mice are randomly divided into groups of control, AR, cineole (30 mg/kg), and budesonide (38.83 µg/kg). The pharmacodynamic results show that cineole significantly reduces the levels of Th2-type cytokines and OVA-specific IgE (OVA-sIgE) in AR mice, improves nasal mucosal tissue damage and alleviates nasal symptoms compared to the untreated AR group. Metabolomic results show that arachidonic acid (AA) metabolism and tryptophan (Trp) metabolism are reprogrammed on the basis of 27 significantly altered metabolites. Further studies show that cineole inhibits the biosynthesis of pro-inflammatory lipid mediators leukotrienes (LTs) and prostaglandins (PGs) in mice by inhibiting the activity of 5-lipoxygenase (5-LOX) and cyclooxygenase-2 (COX-2) in the arachidonic acid metabolic (AA metabolic) pathway. It also inhibits the production of Th2 cytokines and inflammatory cell infiltration, thereby alleviating symptoms such as nasal congestion and nasal leakage. These results reveal the action and molecular mechanism of cineole in alleviating AR and provide a theoretical basis for the clinical application of cineole in treating AR.

Short communication: Antibacterial effects of essential oils from Cinnamomum cassia bark and Eucalyptus globulus leaves-The involvements of major constituents.

Essential oils from Cinnamomum cassia bark and Eucalyptus globulus leaves have been traditionally applied for bacterial infections, through both of aromatherapy and oral application. (E)-cinnamaldehyde and 1,8 cineole have been identified as their major secondary metabolites, and are also generally considered as the main active ingredients responsible for their medicinal applications. However, ethnobotanical doctors still prefer to use whole essentials oils over purified compounds in bacterial infections. We therefore hypothesized that multi-compound extracts might exert better effects than isolated ingredients. In order to verify the hypothesis about advantages of whole materials, we examined antibacterial properties of the 2 plant essential oils in the comparison with their isolated major compounds, such as (E)-cinnamaldehyde and 1,8 cineole. Effects of liquid- and vapor-phase were examined on a set of 6 gram-positive and -negative bacteria, applying broth dilution, agar well diffusion and disc volatilization methods. In all 3 investigations, we observed that whole cinnamon and eucalyptus oils, with the lower concentrations of (E)-cinnamaldehyde (89.1%) and 1,8 cineole (61.2%), were able to induce better effects than the purified active compounds (≥ 99%). These results partly explain the advantages of using whole essential oils over isolated ingredients, and therefore support the application of traditional dosage forms for bacterial infections in ethnomedicine.

Chemical composition, seasonal variation and antiaging activities of essential oil from Callistemon subulatus leaves growing in Egypt.

Callistemon is an aromatic genus of flowering plants belonging to family Myrtaceae. The essential oils of C. subulatus leaves were collected in four seasons and analyzed using GC/MS. The oils demonstrated monoterpenes as the predominant class. Eucalyptol was the main component in all seasons; summer (66.87%), autumn (58.33%), winter (46.74%) and spring (44.63%), followed by α-pinene; spring (31.41%), winter (28.69%), summer (26.34%) and autumn (24.68%). Winter oil, the highest yield (0.53 mL/100g), was further investigated for its inhibitory activity against enzymes associated with ageing; elastase and acetylcholinesterase. It remarkably inhibited elastase and acetylcholinesterase with IC50 values of 1.05 and 0.20 µg/ml, respectively. A molecular docking study was conducted for the major oil components on the active sites of target enzymes. Eucalyptol revealed the best binding affinity for both enzymes. C. subualtus oil could be used as supplement for management of ageing disorders like skin wrinkles and dementia.

Anti-inflammatory mechanisms of eucalyptol rich Eucalyptus globulus essential oil alone and in combination with flurbiprofen.

Inflammation is the core contributor in the pathogenesis of various acute and chronic illness including appendicitis, bronchitis, arthritis, cancer and neurological diseases. NSAIDs, commonly used medications for inflammatory diseases, on prolonged use cause GI bleeding, ulcers and many more issues. Plant-based therapeutic agents including essential oils in combination with low-dose synthetic drugs have been shown to produce synergistic effects and reduce complications of synthetic drugs. This study was designed to evaluate the anti-inflammatory, analgesic and anti-pyretic properties of Eucalyptus globulus essential oil alone and in combination with flurbiprofen. GC-MS analysis was performed to screen chemical composition of oil. In vitro anti-inflammatory assay (membrane stabilization assay) and in vivo inflammatory acute (carrageenan and histamine-induced paw oedema) and chronic (cotton pellet-induced granuloma and Complete Freund's adjuvant-induced arthritis) models were performed to check anti-inflammatory properties. Acetic acid-induced algesia and yeast-induced pyrexia models were performed to check analgesic and anti-pyretic properties. qRT-PCR was performed to study the effect of treatments on the expression of inflammatory biomarkers. GC-MS analysis of E. globulus essential oil showed the presence of eucalyptol along with other active biomolecules. 500 + 10 mg/kg of oil-drug combination showed significantly (p < 0.05) better in vitro membrane stabilization effects as compared with groups treated with 500 mg/kg of E. globulus oil and 10 mg/kg of Flurbiprofen alone. 500 + 10 mg/kg of oil-drug combination showed significantly (p < 0.05) better anti-inflammatory, analgesic and antipyretic effects as compared to 500 mg/kg of E. globulus oil alone in all in vivo models. When comparison was done between 500 + 10 mg/kg of oil-drug combination-treated and 10 mg/kg Flurbiprofen-treated group, the former group showed significantly (p < 0.05) better anti-inflammatory and anti-pyretic effects, but there were non-significant differences in the analgesic model. Animal group treated with 10 mg/kg of Flurbiprofen showed significantly (p < 0.05) better anti-inflammatory and analgesic effects than group treated with 500 mg/kg of oil alone while, there were non-significant differences in anti-pyretic effects. qRT-PCR analysis showed significant (p < 0.05) down-regulation in the expression of IL-4 and TNF-α in serum samples of animals treated with 500 + 10 mg/kg of oil-drug combination as compared to the diseased control (arthritic) group. Overall, the current research demonstrates that Eucalyptus globulus essential oil in combination with flurbiprofen showed better anti-inflammatory, analgesic and anti-pyretic effects than oil and flurbiprofen alone which is attributed to the down-regulation of pro-inflammatory biomarkers (IL-4 and TNF-α). Further studies are required to formulate a stable dosage form and to check the anti-inflammatory efficacy in different inflammatory disorders.

HPTLC Analysis and Chemical Composition of Selected Melaleuca Essential Oils.

Tea tree oil (TTO) is a volatile essential oil obtained by distillation, mainly from the Australian native plant Melaleuca alternifolia (Maiden & Betche) Cheel (Myrtaceae). In this study, a comparative analysis of the chemical constituents of seven tea tree oils (M. alternifolia) and four other Melaleuca spp. oils (M. cajuputi, (MCa), two chemotypes of M. quinquenervia, (MNe and MNi), and M. ericifolia (MRo)) was carried out using gas chromatography-mass spectrometry (GC-MS) and high-performance thin-layer chromatography (HPTLC). Among the seven TTOs, terpinen-4-ol (37.66-44.28%), γ-terpinene (16.42-20.75%), α-terpinene (3.47-12.62%), α-terpineol (3.11-4.66%), and terpinolene (2.75-4.19%) were the most abundant compounds. On the other hand, the most abundant compounds of the other Melaleuca oils varied, such as 1,8-cineole (64.63%) in MCa oil, (E)-nerolidol (48.40%) and linalool (33.30%) in MNe oil, 1,8-cineole (52.20%) in MNi oil, and linalool (38.19%) and 1,8-cineole (27.57%) in MRo oil. HPTLC fingerprinting of Melaleuca oils enabled the discrimination of TTO oils from other Melaleuca spp. oils. Variation was observed in the profile of the Rf values among EOs. The present study shows that HPTLC is one of the best ways to identify and evaluate the quality control in authenticating TTOs, other Melaleuca EOs, or EOs from other species within the Myrtaceae.

Fabrication of a Polysaccharide-Protein/Protein Complex Stabilized Oral Nanoemulsion to Facilitate the Therapeutic Effects of 1,8-Cineole on Atherosclerosis.

Atherosclerosis (AS) is a systemic disease characterized by lipid deposition in the blood vessel wall that urgently requires effective and safe therapeutic drugs for long-term treatment. An essential oil monomer-1,8-cineole (CIN) with ameliorative effects on vascular injuries has considerable potential for preventing the progression of AS because of its antioxidant, anti-inflammation, and cholesterol regulatory effects. However, the high volatility and instability of CIN result in low oral bioavailability and a short half-life, thereby limiting its clinical application. We formulated a nanoemulsion using a polysaccharide-protein/protein complex (dextran-bovine serum albumin/protamine, DEX5k-BSA/PTM) as an emulsifier, with vitamin B12 (VB12) as the ligand to facilitate the transportation across the small intestine. An emulsion preparation method using a microjet followed by ultraviolet irradiation was developed to obtain the CIN-loaded oral nanoemulsion CIN@DEX5k-BSA/PTM/VB12. The nanoemulsion improved the stability of CIN both in vitro and in vivo, prolonged the retention time in the gastrointestinal tract (GIT), and enhanced the permeability across the mucus layer and intestinal epithelial cells to increase oral bioavailability and plaque accumulation of CIN. Validated in an AS mouse model, CIN@DEX5k-BSA/PTM/VB12 achieved prominent therapeutic efficacy combating AS. This study highlights the advantages of DEX5k-BSA/PTM and VB12 in the development of nanoemulsions for CIN and provides a promising oral nanoplatform for the delivery of essential oils.

Evaluation of the yield, chemical composition and biological properties of essential oil from bioreactor-grown cultures of Salvia apiana microshoots.

Microshoot cultures of the North American endemic Salvia apiana were established for the first time and evaluated for essential oil production. Stationary cultures, grown on Schenk-Hildebrandt (SH) medium, supplemented with 0.22 mg/L thidiazuron (TDZ), 2.0 mg/L 6-benzylaminopurine and 3.0% (w/v) sucrose, accumulated 1.27% (v/m dry weight) essential oil, consisting mostly of 1,8-cineole, ß-pinene, α-pinene, ß-myrcene and camphor. The microshoots were adapted to agitated culture, showing biomass yields up to ca. 19 g/L. Scale-up studies demonstrated that S. spiana microshoots grow well in temporary immersion systems (TIS). In the RITA bioreactor, up to 19.27 g/L dry biomass was obtained, containing 1.1% oil with up to ca. 42% cineole content. The other systems employed, i.e. Plantform (TIS) and a custom made spray bioreactor (SGB), yielded ca. 18 and 19 g/L dry weight, respectively. The essential oil content of Plantform and SGB-grown microshoots was comparable to RITA bioreactor, however, the content of cineole was substantially higher (ca. 55%). Oil samples isolated from in vitro material proved to be active in acetylcholinesterase (up to 60.0% inhibition recorded for Plantform-grown microshoots), as well as hyaluronidase and tyrosinase-inhibitory assays (up to 45.8 and 64.5% inhibition observed in the case of the SGB culture).

Chemometrics of the Composition and Antioxidant Capacity of Hyptis crenata Essential Oils from Brazil.

Hyptis crenata (Pohl) ex Benth is used in traditional medicine as an analgesic to treat general pain. Six Hyptis crenata samples (Hc-1 to Hc-6) were collected in Pará state, Brazil. The leaf essential oils were obtained by hydrodistillation, and GC-MS and GC-FID were used to analyze their chemical compositions. The antioxidant capacity was measured in vitro using DPPH and carotene/linoleic acid assays. Chemometrics analysis (PCA, HCA, and clustered heat map) were used to identify the sample relationships between those collected in this study and those from the literature (Hc-7 to Hc-16) samples. According to the main chemical constituents identified in the samples described in this work and the literature, the sixteen samples were classified into ten groups. Group I was characterized by 1,8-cineole (31.0%), α-pinene (13.6%), (E)-caryophyllene (7.8%), and ß-pinene (7.6%); and Group IV was characterized by 1,8-cineole (17.4-23.5%), α-pinene (15.7-23.5%), ß-pinene (10.5-13.4%), and limonene (8.5-9.7%). Both groups are described for the first time. The total antioxidant capacity was expressed in Trolox Equivalent Antioxidant Capacity values (TEAC): TEAC of Hc-5 (551.9 mg.TE/g) and Hc-6 (475.1 mg.TE/g). In the ß-carotene/linoleic acid assay, the highest inhibition was from Hc-2 (40.0%), Hc-6 (39.0%), and Hc-3 (29.4%).

Toxicity and Physiological Effects of Nine Lamiaceae Essential Oils and Their Major Compounds on Reticulitermes dabieshanensis.

The volatile metabolites of Salvia sclarea, Rosmarinus officinalis, Thymus serpyllum, Mentha spicata, Melissa officinalis, Origanum majorana, Mentha piperita, Ocimum basilicum and Lavandula angustifolia were determined by gas chromatography-mass spectrometry. The vapor insecticidal properties of the analyzed essential oils and their compounds were screened using Reticulitermes dabieshanensis workers. The most effective oils were S. sclarea (major constituent linalyl acetate, 65.93%), R. officinalis (1,8-cineole, 45.56%), T. serpyllum (thymol, 33.59%), M. spicata (carvone, 58.68%), M. officinalis (citronellal, 36.99%), O. majorana (1,8-cineole, 62.29%), M. piperita (menthol, 46.04%), O. basilicum (eugenol, 71.08%) and L. angustifolia (linalool, 39.58%), which exhibited LC50 values ranging from 0.036 to 1.670 µL/L. The lowest LC50 values were recorded for eugenol (0.060 µL/L), followed by thymol (0.062 µL/L), carvone (0.074 µL/L), menthol (0.242 µL/L), linalool (0.250 µL/L), citronellal (0.330 µL/L), linalyl acetate (0.712 µL/L) and 1,8-cineole (1.478 µL/L). The increased activity of esterases (ESTs) and glutathione S-transferase (GST) were observed but only alongside the decreased activity of acetylcholinesterase (AChE) in eight main components. Our results indicate that S. sclarea, R. officinalis, T. serpyllum, M. spicata, M. officinalis, O. marjorana, M. piperita, O. basilicum and L. angustifolia essential oils (EOs) and their compounds, linalyl acetate, 1,8-cineole, thymol, carvone, citronellal, menthol, eugenol and linalool could be developed as control agents against termites.

Multivariate Analysis of Essential Oil Composition of Artemisia annua L. Collected from Different Locations in Korea.

Artemisia annua L. is distributed throughout the world and it is an important medicinal plant in Korea to treat various human diseases. Recently, A. annua has also been considered to be an effective ethnobotanical drug against COVID-19. A. annua contains an appreciable amount of essential oil with different biological properties. However, the composition of essential oils in aromatic plants can be varied depending on several factors, including geographic, genetic, ecological, etc. Hence, the present study aimed to investigate the chemical diversity of essential oils of Korean A. annua collected from different locations in Korea by multivariate analysis. For this purpose, the seeds of A. annua were collected from 112 different locations in Korea and were grown under the same environmental conditions. Except for nine individuals which decayed during the cultivation, essential oils were isolated from the aerial parts of 103 A. annua individuals (AEOs) using the steam distillation extraction method, and their chemical compositions were determined by GC-MS analysis. Furthermore, a multivariate analysis was performed to distinguish the difference between 103 individuals of A. annua based on their essential oil compositions. The yield of A. annua essential oils ranged from 0.04 to 1.09% (v/w). Based on the GC-MS data, A. annua individuals were grouped into six chemotypes such as artemisia ketone, camphor, ß-cubebene, eucalyptol, α-pinene, and ß-selinene. The multivariate analysis results revealed that Korean A. annua could be largely grouped into three clusters such as artemisia ketone, eucalyptol, and ß-selinene. Among 35 components selected for principal component analysis (PCA), PC1, PC2, and PC3 accounted for 82.55%, 8.74%, and 3.62%, respectively. Although all individuals of A. annua were cultivated under the same environmental conditions, there is an intraspecific chemical diversity that exists within Korean native species.

The effect of irrigation with treated and untreated wastewater on the yield and chemical composition of essential oil of Mentha spicata L. and Rosmarinus officinalis L.

Today, the lack of quality water supply has led to the tendency to use unconventional water to irrigate agricultural products. Considering the importance and application of essential oils of mint plants in various pharmaceutical, food, and health industries and also considering the approach of using unconventional waters in the cultivation of medicinal plants, the present study aimed to investigate and compare the chemical composition of essential oils of two species of Mentha spicata L. and Rosmarinus officinalis L. which was designed and implemented for the first time under the influence of different treatments of municipal and industrial wastewater. For this purpose, first R. officinalis cuttings and roots of M. spicata were prepared and after preparing and leveling the ground, in the spring of 2020, it was transferred to the planting site and planted in the form of creek and ridges. The treatments studied in this study included well water (WW), treated municipal wastewater (TMW), untreated municipal wastewater (UMW), treated industrial wastewater (TIW), and untreated industrial wastewater (UIW) in a randomized complete block design with four repeat runs. After watering the plants continuously for 3 months, the plant branches were collected and transferred to the laboratory for drying. After extracting the essential oil by water distillation (Clevenger) method, the analysis and identification of the compounds were performed by a chromatograph coupled with a mass spectrometer (GC/MS). The results showed that the highest and lowest yields of M. spicata belonged to the samples treated with UMW and WW, respectively. Also, R. officinalis essential oil irrigated with UMW and UIW had the highest and lowest yields, respectively. The number of essential oil compounds in of M. spicata was between 5 and 19 and in R. officinalis between 14 and 23 under different treatments. The results of the analysis of essential oil compounds showed that D-carvone (57.77-57.44%) and D-limonene (8.70-26.65%) for M. spicata and α-pinene (26.12-34.85%), 1,8-cineole (18.95-23.70%), and camphene (9.93-12.80%) for R. officinalis were predominant compounds in all studied treatments. The results show that UMW is a suitable and efficient treatment to have the best quantity of M. spicata essential oil and the best quality and quantity of R. officinalis essential oil.

Chemical composition and antinociceptive and anti-inflammatory activity of the essential oil of Hyptis crenata Pohl ex Benth. from the Brazilian Amazon.

ETHNOPHARMACOLOGICAL RELEVANCE: The leaf tea of Hyptis crenata has its practical use in the Brazilian Amazon for treating gastrointestinal and liver disorders, sweating induction, and as an anti-inflammatory. AIM OF THE STUDY: Evaluation of the chemical composition, acute oral toxicity, and antinociceptive and anti-inflammatory activities of the H. crenata essential oil. MATERIAL AND METHODS: The essential oil was hydrodistilled and analyzed by GC and GC-MS. The antinociceptive action in mice was evaluated for the peripheral and central analgesic activity (abdominal contortion and hot plate tests), and the xylene-induced ear swelling was carried out for the nociception test. RESULTS: Oxygenated monoterpenes (53.0%) and monoterpene hydrocarbons (38.9%) predominated in the H. crenata oil, being 1,8-cineo1e (35.9%), α-pinene (20.8%), camphor (10.0%), and ß-pinene (7.3%) their primary constituents. The oral oil administration in the mice did not display changes in behavior patterns or animal mortality at 300 and 2000 mg/kg doses. The control group's biochemical parameters (ALP, AST, ALT) displayed a statistical difference from the treated group, unlike the renal parameters, which showed no variation between the groups. Oil reduced the abdominal contortions at doses of 100 (79.5%) and 300 mg/kg (44.4%), while with endodontacin, the dose was 5 mg/kg (75.2%). In addition, the oil could not decrease the paw licking/biting time at doses of 30, 100, and 300 mg/kg. However, it showed a significant antinociceptive effect on the second phase in the formalin test inhibiting licking time, with a reduction of 50.8% (30 mg/kg), 63.4% (100 mg/kg), 58.0% (300 mg/kg), and morphine (4 mg/kg, 78.3%). The oil administration produced significant inhibition of ear edema at all tested doses, with a better effect produced at 30 mg/kg (64.0% inhibition). CONCLUSION: The oil of Hyptis crenata, rich in 1,8-cineole, camphor, α-pinene, and ß-pinene, totaling 74%, displayed low acute toxicity and significant anti-inflammatory activity, with peripheral and no central antinociceptive action. Thus, these results show an actual perspective on using H. crenata oil in developing a phytotherapeutic product.

Thymus mastichina (L.) L. and Cistus ladanifer L. for skin application: chemical characterization and in vitro bioactivity assessment.

ETHNOPHARMACOLOGICAL RELEVANCE: Thymus mastichina (L.) L. (TM) and Cistus ladanifer L. (CL) are two Portuguese autochthonous species with traditional skin application in folk medicine. TM is majorly known for its antiseptic and wound healing properties, as an external anti-inflammatory agent and for its application in folk cosmetics and hygiene products. Its use in acne vulgaris has also been reported. CL is traditionally used in remedies for wounds, ulcers and other skin ailments such as psoriasis and eczema. Its application has been found useful due to its anti-inflammatory, astringent, wound healing and antiseptic properties. AIM OF THE STUDY: With this work, we aimed to investigate relevant bioactivities related with the traditional application of TM and CL essential oils (EOs) and hydrolates (by-products of EO production) in skin ailments. Specifically their in vitro antioxidant, anti-inflammatory, cytotoxic, wound healing and antimicrobial properties were evaluated. The chemical composition of both EOs and respective hydrolates was also characterized. MATERIALS AND METHODS: Chemical characterization of EOs and hydrolates was performed by GC-FID and GC-MS. Cellular biocompatibility was evaluated using the MTT assay in macrophages (RAW 264.7) and fibroblasts (L929) cell lines. Anti-inflammatory activity was investigated by studying nitric oxide (NO) production by macrophages with Griess reagent. Wound healing potential was evaluated with the scratch-wound assay. The antioxidant potential was studied by the DPPH scavenging method. Antimicrobial activity was evaluated by broth microdilution assay against relevant microbial strains and skin pathogens, namely Staphylococcus aureus, Staphylococcus epidermidis, Cutibacterium acnes, Pseudomonas aeruginosa, Escherichia coli, Candida albicans and Aspergillus brasiliensis. RESULTS: The major compounds present in TM and CL EOs were 1,8-cineole and α-pinene, respectively. 1,8-cineole and E-pinocarveol were the major compounds in the correspondent hydrolates. CL EO presented the highest anti-inflammatory potential [EC50 = 0.002% (v/v)], still with significant cytotoxicity [IC50 = 0.012% (v/v)]. TM preparations presented anti-inflammatory potential, also presenting higher biocompatibility. The same profile was present on fibroblasts regarding biocompatibility of the tested preparations. CL EO and hydrolate increased fibroblasts' migration by 155.7% and 148.4%, respectively. TM hydrolate presented a milder activity than CL hydrolate, but wound healing potential was still present, increasing cell migration by 125.1%. All preparations presented poor antioxidant capacity. CL EO presented higher antimicrobial activity, with MICs ranging from 0.06% (v/v) to 2% (v/v), against different microorganisms. CONCLUSIONS: Anti-inflammatory and skin repairing potential were present for CL preparations. TM hydrolate presented an interesting biocompatible profile on both cell lines, also presenting anti-inflammatory potential. Furthermore, EOs from both species presented antimicrobial activity against a panel of different microorganisms. These in vitro bioactivities support some of their traditional skin applications, specifically regarding their antiseptic, wound healing and anti-inflammatory uses.

Quantification of six volatile oil constituents of Oleum Cinnamomi in rat plasma and multiple tissues using GC-MS and its application to pharmacokinetic and tissue distribution studies.

Oleum Cinnamomi is the essential oil obtained from the herb Fructus Cinnamomi which is used by the Hmong people in traditional medicine for the treatment of various diseases. At present, there are a variety of marketed preparations with it as the main medicine on the market. Information regarding the in vivo process of it is lacking, which has become a bottleneck restricting its development and utilization. In view of this, a GC-MS SIM analysis method was established for the simultaneous determination of six main volatile components [eucalyptol, p-cymene, 4-carvomenthenol, 4-isopropyl-2-cyclohexenone, α-terpineol, and 2-(4-Methylphenyl)-propan-2-ol] in plasma and ten tissues of rats to study their pharmacokinetic and distribution characteristics in vivo. The pharmacokinetic results showed that the t1/2 of each index was 0.41-1.66 h, Tmax was 0.16-0.68 h, Cmax was 13.66-2015.02 ng/mL, AUC0-t was 12.84-4299.00 h·ng/mL, CLZ/F was 1750.93-107013.11 mL/h/kg. This meant that the six components could be absorbed quickly, had a short residence time, and be eliminated quickly in the body. Among them, eucalyptol has the highest degree of absorption and a larger amount of entering the body. Moreover, the Cmax and AUC0-t of the six components increased correspondingly with the increase of the dose, indicating that the concentration of Oleum Cinnamomi in the rat plasma was dose-dependent. At different time points, the six components were widely distributed with uneven characteristics in the body. The six components mainly tend to be distributed in stomach, small intestine, and liver, followed by kidney, spleen, heart, and brain, and to a lesser extent in lung, skin, and muscle. And the six components were eliminated quickly in each tissue. The pharmacokinetic process and tissue distribution characteristics of Oleum Cinnamomi were expounded in this study, which can provide scientific theory for the in-depth development and guidance of clinical drug use of Oleum Cinnamomi, and at the same time provide a medicinal material basis for the in-depth development and utilization of Oleum Cinnamomi.

Treatment with Glyphosate Induces Tolerance of Citrus Pathogens to Glyphosate and Fungicides but Not to 1,8-Cineole.

During the postharvest period, citrus fruits are exposed to Penicillium italicum, Penicillium digitatum, and Geotrichum candidum. Pesticides such as imazalil (IMZ), thiabendazole (TBZ), orthophenylphenol (OPP), and guazatine (GUA) are commonly used as antifungals. Glyphosate (GP) is also used in citrus fields to eliminate weed growth. The sensitivity of fungal pathogens of citrus fruit to these pesticides and 1,8-cineole was evaluated, and the effect of GP on the development of cross-resistance to other chemicals was monitored over a period of 3 weeks. IMZ most effectively inhibited the mycelial growth and spore germination of P. digitatum and P. italicum, with minimum inhibitory concentrations (MICs) of 0.01 and 0.05 mg/mL, respectively, followed by 1,8-cineole, GP, and TBZ. 1,8-Cineole and GP more effectively inhibited the mycelial growth and spore germination of G. candidum, with minimum inhibitory concentrations (MICs) of 0.2 and 1.0 mg/mL, respectively, than OPP or GUA. For the spore germination assay, all substances tested showed a total inhibitory effect. Subculturing the fungal strains in culture media containing increasing concentrations of GP induced fungal tolerance to GP as well as to the fungicides. In soil, experiments confirmed that GP induced the tolerance of P. digitatum to TBZ and GP and the tolerance of P. italicum to IMZ, TBZ, and GP. However, no tolerance was recorded against 1,8-cineole. In conclusion, it can be said that 1,8-cineole may be recommended as an alternative to conventional fungicides. In addition, these results indicate that caution should be taken when using GP in citrus fields.

Transcriptome Analysis Reveals the Anti-Tumor Mechanism of Eucalyptol Treatment on Neuroblastoma Cell Line SH-SY5Y.

Eucalyptol (1.8-cineole), an active component in traditional Chinese medicine Artemisia argyi for moxibustion. Previous studies have shown that eucalyptol has anti-tumor effects on leukemia and colon cancer. Nonetheless, the effect and mechanism of eucalyptol on neuroblastoma remains unclear. In the present study, we intended to reveal the effect and mechanism of eucalyptol treatment on the neuroblastoma cell line SH-SY5Y through transcriptome analysis. In the group treated with eucalyptol, 566 brain genes were up-regulated, while 757 genes were down-regulated. GO function analysis showed that positive regulation of cell cycle was down-regulated in biological processes. Meanwhile, cancer-related pathways were identified in KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment analysis, including pathways in cancer, PI3K-Akt signaling pathway, cAMP signaling pathway, TGF-beta signaling pathway, Hippo signaling pathway, p53 signaling pathway, and additional pathways. Furthermore, we found a key gene, such as MYC, by constructing a network of cancer related pathways with differentially expressed genes and transcription factor analysis. In conclusion, our research indicates that MYC might play a central role in the anit-tumor mechanisms of eucalyptol.