Chemical Variability of the Berry Essential Oil of Two Subspecies of Juniperus Oxycedrus L. from Northwestern Algeria.

The chemical composition of 33 oil samples isolated from the berries of individual trees of two subspecies of Juniperus oxycedrus L., collected in four locations in Tlemcen and Aïn Temouchent provinces, was investigated by GC-FID, GC/MS and 13CNMR. In total, 83 compounds were identified, accounting for 85.9 % to 98.6 % of the total composition. Yields ranged between 0.02 and 0.40 % (w/w). All compositions were submitted to statistical analysis. Combination of hierarchical clustering dendrogram and principal component analysis suggested the existence of two groups, the second group being sub-divided into two sub-groups. Groups and sub-groups were distinguished with respect to their contents of α-pinene, germacrene D, myrcene and, to a lesser extent, of manoyl oxide. The chemical composition of the oils belonging to subsp. oxycedrus was largely dominated by monoterpenes. α-Pinene (23.7-63.2 %) and germacrene D (4.5-25.4 %) were the major components, followed by myrcene (3.1-16.7 %). Conversely, samples oils of subsp. macrocarpa were largely dominated by germacrene D (4.4-30.0 %), followed by a lower content of α-pinene (10.0-24.3 %) and myrcene (1.0-4.3 %) and a significant content of manoyl oxide (4.3-9.9 %).

Antispasmodic and antidiarrheal effects of Juniperus oxycedrus L. on the jejunum in rodents.

Functional bowel disorders (FBD) have a major potential to degrade the standards of public life. Juniperus oxycedrus L. (J. oxycedrus) (Cupressaceae) has been described as a plant used in traditional medicine as an antidiarrheal medication. The present study is the first to obtain information on the antispasmodic and antidiarrheic effects of J. oxycedrus aqueous extract through in vitro and in vivo studies. An aqueous extract of J. oxycedrus (AEJO) was extracted by decoctioning air-dried aerial sections of the plant. Antispasmodic activity was tested in an isolated jejunum segment of rats exposed to cumulative doses of drogue extract. The antidiarrheic activity was tested using diarrhea caused by castor oil, a transit study of the small intestine, and castor oil-induced enteropooling assays in mice. In the jejunum of rats, the AEJO (0.1, 0.3 and 1 mg/ml) diminished the maximum tone induced by low K+ (25 mM), while it exhibited a weak inhibitory effect on high K+ (75 mM) with an IC50=0.49 ± 0.01 mg/ml and IC50=2.65 ± 0.16 mg/ml, respectively. In the contractions induced by CCh (10-6 M), AEJO diminished the maximum tone, similar to that induced by low K+ (25 mM). with an IC50=0.45 ± 0.02 mg/ml. The inhibitory effect of AEJO on low K+ induced contractions was significantly diminished in the presence of glibenclamide (GB) (0.3 µM) and 4-aminopyrimidine (4-AP) (100 µM), with IC50 values of 1.84 ± 0.09 mg/ml. and 1.63 ± 0.16 mg/ml, respectively). The demonstrated inhibitory effect was similar to that produced by a non-competitive antagonist acting on cholinergic receptors and calcium channels. In castor oil-induced diarrhea in mice, AEJO (100, 200, and 400 mg/kg) caused an extension of the latency time, a reduced defecation frequency, and a decrease in the amount of wet feces compared to the untreated group (distilled water). Moreover, it showed a significant anti-motility effect and reduced the amount of fluid accumulated in the intestinal lumen at all tested doses. These findings support the conventional use of Juniperus oxycedrus L. as a remedy for gastrointestinal diseases.

Phytochemical Analysis and Evaluation of Antifungal and Antioxidant Activities of Essential Oil of Fruits from Juniperus oxycedrus L. Obtained from Morocco

Abstract The present study was aimed at conducting phytochemical analysis and evaluating the in vitro antifungal and antioxidant activities of the essential oil obtained from the fruits of J. oxycedrus L. Hydro-distillation was used to extract the essential oil from the fruits of Juniper oxycedrus. The essential oil was analyzed using gas chromatography with a flame ionization detector (GC-FID) and gas chromatography coupled with mass spectrometry (GC/MS). The antioxidant activity of the essential oil against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals was determined in vitro using varying concentrations of the essential oil and vitamin C as a standard antioxidant compound. A disc diffusion test was employed to evaluate the antifungal activity of the essential oil against two test fungal strains, Penicillium citrinum, and Aspergillus niger. The results revealed that 49 constituents were identified in fruit oil, representing 91.56% of the total oil and the yield was 1.58%. Juniper fruit oil was characterized by having high contents of ß-pinene (42.04%), followed by limonene (15.45%), sabinene (9.52%), α-pinene (5.21%), (E)-caryophyllene (3.77%), ρ-cymene (1.56%), caryophyllene oxide (2.02%), and myrcene (1.02%). The radical scavenging activity (% inhibition) of the essential oil was highest (81.87± 2.83%) at a concentration of 200 µg/mL. The essential oil of J. oxycedrus exhibited antifungal activity against A. niger and P. citrinum with minimum inhibitory concentration values (MIC) ranging from 2.89 to 85.01 µl/mL. The findings of the study reveal that the antioxidant and antifungal properties of J. oxycedrus essential oil and their chemical composition are significantly correlated

Evaluation of the Antimicrobial and Antivirulent Potential of Essential Oils Isolated from Juniperus oxycedrus L. ssp. macrocarpa Aerial Parts.

As a consequence of the worsening situation with multidrug-resistant (MDR) pathogens and a disparity in the commercialization of novel antimicrobial agents, scientists have been prompted to seek out new compounds with antimicrobial activity from a wide range of sources, including medicinal plants. In the present study, the antibacterial, antifungal, anti-virulence, and resistance-modulating properties of the essential oil from the Sardinian endemic Juniperus oxycedrus L. ssp. macrocarpa aerial parts were evaluated. The GC/MS analysis showed that the main compounds in the oil were α-pinene (56.63 ± 0.24%), limonene (14.66 ± 0.11%), and ß-pinene (13.42 ± 0.09%). The essential oil showed potent antibacterial activity against Gram-positive bacteria (0.25-2 v/v%) and Salmonella spp. (4 v/v%). The strongest fungicidal activity was recorded against Candida auris sessile cells (median FICI was 0.088) but not against C. albicans biofilms (median FICI was 1). The oil showed potent efflux pump inhibitory properties in the case of Staphylococcus aureus and Escherichia coli. The therapeutic potential of Juniperus may be promising for future more extensive research and in vivo tests to develop new drugs against antibiotic and antifungal resistance.

Isolation and characterization of a mesophilic cellulolytic endophyte Preussia africana from Juniperus oxycedrus.

The medicinal plant Juniperus oxycedrus is less recognized for the diversity of its fungal endophytes and their potential to produce extracellular enzymes. The present study is the first report on the isolation and identification of a mesophilic endophytic strain JO-A, Preussia africana, from fresh stems of the J. oxycedrus endemic tree in the Ifrane region-Morocco, and the evaluation of its ability to produce cellulases. A one-time multi-parameter one-factor screening was optimized to select factors that enhance cellulase production in P. africana. The maximum production of both CMCase and FPase activities were 1.913 IU.mL-1 and 0.885 IU.mL-1, respectively, when the medium was supplemented with 2% w/v glucose. These remarkable titers were tenfold greater than those obtained under the initial non-optimized conditions. This mesophilic P. africana JO-A strain grows and actively produces cellulases at 37 °C demonstrating its great potential for various biotechnology applications. The cellulolytic extract showed the highest enzymatic activities at pH 5.0 and 50 °C with a half-life of 24 h at 50 °C.

Survival, growth, and biogenic amine production of Enterococcus faecium FC12 in response to extracts and essential oils of Rubus fruticosus and Juniperus oxycedrus.

Introduction: Enterococci are lactic acid bacteria (LAB) usually found as food contaminants in fermented products such as cheeses and fermented sausages. Due to their antibiotic resistance, the presence of virulence factors, and the ability to produce biogenic amines (BAs), the determination of these bacteria is crucial to assure food quality and safety. BAs production and consequent accumulation in foods can cause toxicological effects on human health. Plant phenolic compounds are promising alternatives to chemical preservatives and reflect consumers' demand for "green" solutions. In this study, the antimicrobial effect of blackberry (Rubus fruticosus) leaves and prickly juniper (Juniperus oxycedrus) needles, both as phenolic extracts (PE) and essential oils (EO), were evaluated against Enterococcus faecium FC12, a known tyramine-producing strain. Methods: The growth kinetics in the presence of sub-lethal concentrations of such plant derivatives were modeled (Gompertz equation) and BA production was monitored over time by HPLC. Moreover, flow cytometry (FCM) was used to study the effects of EOs and PEs on cell viability. Results: The EOs showed a higher antimicrobial effect (especially R. fruticosus added at 0.75 mg/ml), determining an initial decrease of culturable cells followed by a recovery, even if with lower growth rates and final cell loads. Different rates of BA formation were observed, with tyramine concentrations ranging from 120 to 160 mg/l after 96 h of incubation, and 2-phenylethylamine was produced in lower amounts, usually after reaching the peak of tyramine. FCM confirmed the higher efficacy of R. fruticosus EO that induced cell membrane injury in 93% of the total population. However, complete recovery occurred in the following incubation, demonstrating transient damage. Discussion: Although further research is required to better investigate this recovery and to assess the suitability of this approach in a real food system, the present study showed the potential antimicrobial activity of plant derivatives, especially R. fruticosus EO, against the tyramine-producing E. faecium FC12.

Essential oil constituents of Juniperus oxycedrus L. and Cupressus sempervirens L. (Cupressaceae) growing in Aures region of Algeria.

The purpose of this study was to determine the chemical composition of essential oils from the aerial parts of two species belonging to the Cupressaceae family growing in the Aures region of Algeria Juniperus oxycedrus and Cupressus Sempervirens. The analysis by GC-MS and GC-FID techniques showed the presence of 38 compounds in J. oxycedrus oil where the major constituents were manoyl oxide (23.5%), pentadecan-2-enone 6Z (12.6%), abietatriene (8.0%), abieta-8,11,13-triene-7-one (6.5%), cubebol (4.6%), epi-torilenol (3.8%) and α-cadinol (2.6%), while, a total of 65 compounds were showed in C. sempervirens oil where the major constituents were α-pinene (68.0%), epi-cedrol (6.1%), α-terpenyl acetate (3.5%) and germacrene D (2.5%). It is the first time that the compounds pentadecan-2-enone 6Z, abieta-8,11,13-trien-7-one, cubebol and epi-torilenol have been identified in our J. oxycedrus and epi-cedrol in C. sempervirens essential oils with high contents, as we noticed the absence of α-pinene in our J. oxycedrus essential oil.

Chemical Composition, Antioxidant, Anti-Tyrosinase, Anti-Cholinesterase and Cytotoxic Activities of Essential Oils of Six Algerian Plants.

In this study, the essential oils (EOs) of six Algerian plants (Artemisia campestris L., Artemisia herba-alba Asso, Juniperus phoenicea L., Juniperus oxycedrus L., Mentha pulegium L. and Lavandula officinalis Chaix) were obtained by hydrodistillation, and their compositions determined by GC-MS and GC-FID. The antioxidant activity of the EOS was evaluated via 2,2'-diphenyl-1-picrylhydrazyl (DPPH), ferric-reducing/antioxidant power (FRAP) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS) assays. Moreover, their cytotoxic effect was evaluated-as well as their tyrosinase, acetyl- and butyryl-cholinesterase (AChE and BuChE) inhibitory activities. The chemical analyses detected 44, 45, 51, 53, 26 and 40 compounds in EOs of A. campestris, A. herba-alba, J. phoenicea, J. oxycedrus, M. pulegium and L. officinalis, respectively. A. campestris EO was mainly composed of ß-pinene (20.7%), while A. herba-alba EO contained davanone D (49.5%) as the main component. α-Pinene (41.8%) was detected as the major constituent in both J. phoenicea (41.8%) and J. oxycedrus (37.8%) EOs. M. pulegium EO was characterized by pulegone as the most abundant (76.9%) compound, while linalool (35.8%) was detected as a major constituent in L. officinalis EO. The antioxidant power evaluation revealed IC50 values ranging from 2.61 to 91.25 mg/mL for DPPH scavenging activity, while the FRAP values ranged from 0.97-8.17 µmol Trolox equivalents (TX)/g sample. In the ABTS assay, the values ranged from 7.01 to 2.40 µmol TX/g sample. In the presence of 1 mg/mL of the samples, tyrosinase inhibition rates ranged from 11.35% to 39.65%, AChE inhibition rates ranged from 40.57% to 73.60% and BuChE inhibition rates ranged from 6.47% to 72.03%. A significant cytotoxic effect was found for A. herba-alba EO. The obtained results support some of the traditional uses of these species in food preservation and for protection against several diseases.

Phytochemical Characterization, Antioxidant and In Vitro Cytotoxic Activity Evaluation of Juniperus oxycedrus Subsp. oxycedrus Needles and Berries.

In order to evaluate the antioxidant properties of aqueous and methanol extracts of needles and berries of Juniperus oxycedrus subsp. oxycedrus (Joo) species, various antioxidant capacity assessment tests (free radical scavenging assays (DPPH• and ABTS•+ tests), ferrous ions (Fe2+) chelating activity and reducing power assay (FRAP) were conducted. In all of the tests, the extracts exhibited strong antioxidant activity. Furthermore, in-vitro cytotoxic activity assays of the methanolic extracts showed potent cytotoxic effects against two breast cancer cell lines (MDA-MB-468 and MCF-7), with no cytotoxicity towards normal cells (PBMCs). Reactive oxygen species generation was presumed to be a potential reason for the observed cytotoxic effects. According to all the above, and considering its appropriate composition of mineral elements and phenolic compounds, Joo could offer a beneficial and natural source of bioactive compounds that can be either used on the preventive side as it could potentially be used in the clinic without toxicity.

In Vitro Antimicrobial Activity of Essential Oils from Sardinian Flora against Cutibacterium (Formerly Propionibacterium) acnes and Its Enhancement by Chitosan.

The in vitro antibacterial activity of some essential oils from Sardinian flora, both alone and in combination with chitosan, was investigated against a strain of Cutibacterium acnes, a bacterium involved in pathogenesis of acne. The composition of the essential oils was determined by gas chromatography and gas chromatography/mass spectrometry. The results of this investigation demonstrated that some of the oils examined, characterised by different chemical profiles, possessed some activity against C. acnes. Interestingly, this antibacterial effect was enhanced by sub-inhibitory concentrations of chitosan. These observations suggest the potential application of this synergy in the development of innovative topical formulations useful in the management of acne.

In vitro susceptibility of Trypanosoma brucei brucei to selected essential oils and their major components.

Aiming for discovering effective and harmless antitrypanosomal agents, 17 essential oils and nine major components were screened for their effects on T. b. brucei. The essential oils were obtained by hydrodistillation from fresh plant material and analyzed by GC and GC-MS. The trypanocidal activity was assessed using blood stream trypomastigotes cultures of T. b. brucei and the colorimetric resazurin method. The MTT test was used to assess the cytotoxicity of essential oils on macrophage cells and Selectivity Indexes were calculated. Of the 17 essential oils screened three showed high trypanocidal activity (IC50 < 10 µg/mL): Juniperus oxycedrus (IC50 of 0.9 µg/mL), Cymbopogon citratus (IC50 of 3.2 µg/mL) and Lavandula luisieri (IC50 of 5.7 µg/mL). These oils had no cytotoxic effects on macrophage cells showing the highest values of Selectivity Index (63.4, 9.0 and 11.8, respectively). The oils of Distichoselinum tenuifolium, Lavandula viridis, Origanum virens, Seseli tortuosom, Syzygium aromaticum, and Thymbra capitata also exhibited activity (IC50 of 10-25 µg/mL) but showed cytotoxicity on macrophages. Of the nine compounds tested, α-pinene (IC50 of 2.9 µg/mL) and citral (IC50 of 18.9 µg/mL) exhibited the highest anti-trypanosomal activities. Citral is likely the active component of C. citratus and α-pinene is responsible for the antitrypanosomal effects of J. oxycedrus. The present work leads us to propose the J. oxycedrus, C. citratus and L. luisieri oils as valuable sources of new molecules for African Sleeping Sickness treatment.

Toxicity of Juniperus oxycedrus oil constituents and related compounds and the efficacy of oil spray formulations to Dermatophagoides farinae (Acari: Pyroglyphidae).

The American house dust mite (AHDM), Dermatophagoides farinae Hughes (Acari: Pyroglyphidae), is recognized as an important source of allergens in the domestic environment. This study was conducted to determine whether 19 constituents from essential oil of cade, Juniperus oxycedrus L. (Cupressaceae), eight structurally related compounds, and another 16 previously known cade oil constituents were toxic for adult AHDMs and to determine the route of acaricidal action of the test compounds, as well as to assess the control efficacy of four experimental spray formulations containing the oil (10-40 mg/L sprays). In a fabric-circle contact mortality bioassay, methyleugenol (LD50, 5.82 µg/cm2) and guaiacol (8.24 µg/cm2) were the most toxic compounds against the mites, and the toxicity of these compounds and benzyl benzoate did not significantly differ. High toxicity was also observed with eugenol, m-cresol, and nerolidol (LD50, 12.52-19.52 µg/cm2), and these compounds were significantly more toxic than N,N-diethyl-3-methylbenzamide (DEET) (LD50, 37.67 µg/cm2). Cade applied as 30 or 40 mg/L experimental sprays provided 96 and 100% mortality against the mites, respectively, whereas permethrin (cis:trans, 25:75) 2.5 g/L spray treatment resulted in 17% mortality. In vapor-phase mortality tests, the compounds described were consistently more toxic in closed versus open containers, indicating that toxicity was achieved mainly through the action of vapor. Reasonable mite control in indoor environments can be achieved by spray formulation containing the 40 mg/L cade oil as potential contact-action fumigants.

Taxonomy of prickly juniper (Juniperus oxycedrus group): A phytochemical-morphometric combined approach at the contact zone of two cryptospecies.

Based on different essential oil composition paralleling different genotypes, Juniperus deltoides was recently segregated from Juniperus oxycedrus. Despite a clear phytochemical and molecular differentiation, J. deltoides resulted not clearly morphologically discernible from J. oxycedrus, so that it was defined as a cryptospecies. Italy represents the contact zone of their distribution, but the ranges of the two species are not sufficiently known, due to unsatisfactory morphological characterisation. To further complicate the picture, a third closely related species (ecotype), J. macrocarpa, occurs all across the Mediterranean coasts. After a preliminary phytochemical analysis to ascertain the (chemo-)identities of the studied populations, we performed a morphometric investigation to test the degree of morphological distinctiveness among the taxa. According to our analysis, some character (e.g. leaf mucro length, leaf width, seed-cone size and seed size) resulted useful to discriminate these cryptic taxa. Finally, based on these characters, an extensive revision of herbarium specimens allowed us to redefine the distribution pattern of the investigated species in the Central Mediterranean area.

Side effects of cade oil in Morocco: an analysis of reports in the Moroccan herbal products database from 2004 to 2012.

Cade oil is a dark, faintly aromatic oil which is distilled from the branches and wood of Juniperus oxycedrus. Although this oil is known to have toxic effects related to its content of phenols, cade oil continues to be used in folk medicine. Because of this use, a determination of the safety and possible side effects of cade oil is required. The safety of cade oil is discussed based on the experience of the Moroccan pharmacovigilance herbal products database, and supported by literature. The data on the adverse effects of cade oil suggests that it could have life-threatening effects which can occur following topical exposure, ingestion or inhalation. Phenol's adverse effects involve a wide variety of organ systems such as the gastro-intestinal system, central and peripheral nervous systems, cardiovascular, liver and biliary systems, the urinary tract, skin and appendages, respiratory system. Platelet function, bleeding and clotting, vision, metabolism, and white cell and reticuloendothelial system function are also affected.

Juniperus oxycedrus L. subsp. oxycedrus and Juniperus oxycedrus L. subsp. macrocarpa (Sibth. & Sm.) Ball. "berries" from Turkey: comparative evaluation of phenolic profile, antioxidant, cytotoxic and antimicrobial activities.

This work aimed to evaluate and compare the phenolic profile and some biological properties of the ripe "berries" methanol extracts of Juniperus oxycedrus L. subsp. oxycedrus (Joo) and Juniperus oxycedrus L. subsp. macrocarpa (Sibth. & Sm.) Ball. (Jom) from Turkey. The total phenolic content resulted about 3-fold higher in Jom (17.89±0.23 mg GAE/g extract) than in Joo (5.14±0.06 mg GAE/g extract). The HPLC-DAD-ESI-MS analysis revealed a similar flavonoid fingerprint in Joo and Jom, whereas a difference in their quantitative content was found (4632 µg/g extract and 12644 µg/g extract). In addition, three phenolic acids were detected in Jom only (5765 µg/g extract), and protocatechuic acid was the most abundant one. The antioxidant capacity of the extracts was evaluated by different in vitro assays: in the DPPH and in the TBA tests a stronger activity in Jom was highlighted, while Joo exhibited higher reducing power and metal chelating activity. Joo and Jom did not affect HepG2 cell viability and both extracts resulted virtually non-toxic against Artemia salina. The extracts were also studied for their antimicrobial potential, displaying efficacy against Gram-positive bacteria.