Browsing by Author "Esteban Pascual, Luis Saul"
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- Antioxidant activity and GC-MS characterization of juniperus communis L. and cistus ladanifer L. essential oilsPublication . Xavier, Virginie; Heleno, Sandrina A.; Prieto Lage, Miguel A.; Amaral, Joana S.; Mediavilla Ruiz, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianJuniperus communis L. and Cistus ladanifer L. are two abundant shrubs in the mountain areas of the Mediterranean basin, particularly in the Iberian Peninsula. Both species are known for their valuable essential oil that can be used in cosmetic, food and pharmaceutical industries for their bioactive properties [1]. Within the scope of the European Project “BeonNat”, that aims at developing innovative and bio-based products using the biomass of trees and shrubs growing on marginal and underutilised lands as feedstock for the bio-based industry, these two species were selected for evaluation. In this work, the biomass of the two species (branches milled to <20mm particle size) grown in Spain was submitted to steam distillation . The extracted essential oils (EO) were characterized regarding their chemical composition by gas chromatography coupled with mass spectrometry (GC-MS) and their antioxidant activity by the ferric reducing power assay. An extraction yield of 0.50% and 0.08% (dry basis) was obtained for J. communis and C. ladanifer biomasses, respectively. The GC-MS analysis enabled the identification of 98.1% of compounds in J. communis EO, corresponding to a total of 63 identified compounds, with α-pinene being the major compound (32.3%), followed by limonene (15.8%), sabinene (7.6%), germacrene B (4.9%), cis- t hujopsene (4.6%), β-myrcene (3.7%) and β-caryophyllene (3.6%). In general, the chemical composition was in good agreement with that of juniper berries essential oil defined in the European Pharmacopoeia and the ISO 8897 standard, with the exception of limonene (15.8%) that was slightly higher than the defined range (Eur. Ph from 2-12% and ISO standard from 2-8%). For C. ladanifer essential oil, a total of 61 compounds were identified corresponding to 92.8% of total compounds, with viridiflorol being the major compound (20.7%), followed by α-pinene (19.8%), ledol (8.1%), camphene (7.2%) and bornyl acetate (5.6%). This result is in good agreement with previous works that also report the sesquiterpene alcohol viridiflorol as the major compound in the EO obtained from C. ladanifer leaves and small branches [2]. Regarding the antioxidant activity, both oils showed promising results in the reducing power assay, presenting an EC 50 value of 1.35 ± 0.19 mg/mL and 1.30 ± 0.07 mg/mL for J. communis and C. ladanifer , respectively.
- Assessment of the use of common juniper (Juniperus communis L.) foliage following the cascade principlePublication . Mediavilla, Irene; Bados, Raquel; Barros, Lillian; Xavier, Virginie; Finimundy, Tiane C.; Pires, Tânia C.S.P.; Heleno, Sandrina A.; Calhelha, Ricardo C.; Amaral, Joana S.; Rizzo, Andrea Maria; Casini, David; Lombardi, Giacomo; Chiaramonti, David; Camara, Miguel; Suarez, Ana; Ardid, Tomas; Esteban Pascual, Luis SaulJuniperus communis L. is a species commonly grown in regions of the Northern Hemisphere, and is a good candidate to be cultivated in marginal lands. Plants coming from a pruning performed in a natural population located in Spain were used to assess the yield and quality of different products obtained following the cascade principle. A total of 1050 kg of foliage biomass were crushed, steam-distilled, and separated into fractions to produce biochar and absorbents for the pet industry using pilot plants. The obtained products were analysed. The essential oil, with a yield of 0.45% dry basis and a qualitative chemical composition similar to that described for the berries in international standards or monographs, showed antioxidant activity with promising CAA results (inhibition of 89% of the cell's oxidation). However, regarding antibacterial and antifungal activities, it only inhibited the growth of microorganisms at the maximum concentration tested, 2.5%. Concerning the hydrolate, it did not show bioactivity. Regarding the biochar, whose yield was 28.79% dry basis, interesting results were obtained for its characterisation as a possible soil improver for agronomic purposes (PFC 3(A)). Finally, promising results were obtained regarding the use of common juniper as absorbent, taking into account the physical characterisation and odour control capacity.
- Bioactivities and GC-MS characterization of cupressus sempervirens L. and rosmarinus officinalis L. essential oilsPublication . Xavier, Virginie; Heleno, Sandrina A.; Prieto Lage, Miguel A.; Amaral, Joana S.; Mandim, Filipa; Vaz, Josiana A.; Pires, Tânia C.S.; Mediavilla Ruiz, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianBio-based products will play an increasingly important role in the transition towards a green and circular economy that will reduce fossil-based products consumption. 1 The European Project “BeonNat” aims at developing different bio-based products from underutilised shrubs and trees, which will be used as raw-materials for the bio-based industries. Within this scope, Cupressus sempervirens L. and Rosmarinus officinalis L. were selected for evaluation. In this work, the essential oil of C. sempervirens (common cypress) and R. officinalis (rosemary) cultivated in Spain, were characterized for their chemical composition and potential biological properties. The assessment of antioxidant, antimicrobial, cytotoxic and anti-inflammatory properties of the essential oils were evaluated. The GC-MS analysis enabled the identification of 85% of total compounds in common cypress and 94% in rosemary. The major compounds were α-pinene, 3-carene, cedrol, limonene and terpinolene for C. sempervirens and α-pinene, 1,8-cineole, camphor, camphene and β-myrcene for R. officinalis essential oils. From a qualitative point of view, the obtained results are in good agreement with the data available from the literature, considering the genetics and environment variations that may occur. 2,3 Both samples showed antibacterial activity against a panel of bacteria selected according to their importance in public health, highlighting R. officinalis that revealed the strongest activity for Escherichia coli, Klebsiella pneumoniae, Morganella morganii, Pseudomonas aeruginosa, Enterococcus faecalis, Listeria monocytogenes and methicillin resistant Staphylococcus aureus. Concerning the antioxidant activity, reducing power and cellular antioxidant assays were performed. The cypress essential oil presented an EC50 value of 1.518±0.004 mg/mL using reducing power assay and a % of oxidation inhibition of 73% with cellular antioxidant assay, while rosemary essential oil showed an EC50 value of 3.12±0.01 mg/mL using reducing power assay and 84 % of oxidation inhibition according to cellular antioxidant assay, evidencing that both samples revealed capacity to inhibit the oxidation process. The tested essential oils showed anti-inflammatory (IC50 value of 11±1 μg/mL and 20±1 μg/mL for C. sempervirens and R. officinalis, respectively) and cytotoxic activity, with best results being obtained for C. sempervirens essential oil on the inhibition of breast, MCF-7, (GI50 value of 62±6 μg/mL) and lung cancer, NCI-H460, (GI50 value of 14±1 μg/mL) cell lines. After screening the essential oils for their bioactivities, the results revealed that essential oils from common cypress and rosemary are a potential and natural source of bioactive substances to use in food, cosmetic and medicinal industry, encouraging the valorisation of underexplored resources.
- Bioactivities and GC-MS characterization of Cupressus Sempervirens L. and Rosmarinus Officinalis L. essential oilsPublication . Xavier, Virginie; Heleno, Sandrina A.; Prieto Lage, Miguel A.; Amaral, Joana S.; Mandim, Filipa; Vaz, Josiana A.; Pires, Tânia C.S.; Mediavilla Ruiz, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianBio-based products will play an increasingly important role in the transition towards a green and circular economy that will reduce fossil-based products consumption. 1 The European Project “BeonNat” aims at developing different bio-based products from underutilised shrubs and trees, which will be used as raw-materials for the bio-based industries. Within this scope, Cupressus sempervirens L. and Rosmarinus officinalis L. were selected for evaluation. In this work, the essential oil of C. sempervirens (common cypress) and R. officinalis (rosemary) cultivated in Spain, were characterized for their chemical composition and potential biological properties. The assessment of antioxidant, antimicrobial, cytotoxic and anti-inflammatory properties of the essential oils were evaluated. The GC-MS analysis enabled the identification of 85% of total compounds in common cypress and 94% in rosemary. The major compounds were α-pinene, 3-carene, cedrol, limonene and terpinolene for C. sempervirens and α-pinene, 1,8-cineole, camphor, camphene and β-myrcene for R. officinalis essential oils. From a qualitative point of view, the obtained results are in good agreement with the data available from the literature, considering the genetics and environment variations that may occur. 2,3 Both samples showed antibacterial activity against a panel of bacteria selected according to their importance in public health, highlighting R. officinalis that revealed the strongest activity for Escherichia coli, Klebsiella pneumoniae, Morganella morganii, Pseudomonas aeruginosa, Enterococcus faecalis, Listeria monocytogenes and methicillin resistant Staphylococcus aureus. Concerning the antioxidant activity, reducing power and cellular antioxidant assays were performed. The cypress essential oil presented an EC50 value of 1.518±0.004 mg/mL using reducing power assay and a % of oxidation inhibition of 73% with cellular antioxidant assay, while rosemary essential oil showed an EC50 value of 3.12±0.01 mg/mL using reducing power assay and 84 % of oxidation inhibition according to cellular antioxidant assay, evidencing that both samples revealed capacity to inhibit the oxidation process. The tested essential oils showed anti-inflammatory (IC50 value of 11±1 μg/mL and 20±1 μg/mL for C. sempervirens and R. officinalis, respectively) and cytotoxic activity, with best results being obtained for C. sempervirens essential oil on the inhibition of breast, MCF-7, (GI50 value of 62±6 μg/mL) and lung cancer, NCI-H460, (GI50 value of 14±1 μg/mL) cell lines. After screening the essential oils for their bioactivities, the results revealed that essential oils from common cypress and rosemary are a potential and natural source of bioactive substances to use in food, cosmetic and medicinal industry, encouraging the valorisation of underexplored resources.
- Chemical and bioactive characterization of the essential oils obtained from three mediterranean plantsPublication . Xavier, Virginie; Finimundy, Tiane C.; Heleno, Sandrina A.; Amaral, Joana S.; Calhelha, Ricardo C.; Vaz, Josiana A.; Pires, Tânia C.S.; Mediavilla, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianCupressus sempervirens L., Juniperus communis L. and Cistus ladanifer L. are Mediterranean arboreal and shrub species that possess essential oils (EO) in their leaves and branches. This study aimed at characterizing the EOs obtained by steam distillation from the three species collected in different locations from Spain (Almazán, Andévalo, Barriomartín, Cerezal, Ermitas and Huéscar). For this purpose, volatiles composition was determined by GC-MS, and different bioactivities were evaluated. The highest content in terpenes was observed in C. sempervirens (Huéscar origin) followed by J. communis (Almazán origin), corresponding to 92% and 91.9% of total compounds, respectively. With exception of C. ladanifer from Cerezal that presented viridiflorol as the most abundant compound, all the three species presented in common the -pinene as the major compound. The EOs from C. ladanifer showed high antibacterial potential, presenting MIC values from 0.3 to 1.25 mg/mL. Concerning other bioactivities, C. ladanifer EO revealed an oxidation inhibition of 83%, while J. communis showed cytotoxicity in the MCF-7 cell line, and C. sempervirens and C. ladanifer EOs exhibited the highest potential on NCI-H460 cell lines. Nevertheless, some EOs revealed toxicity against non-tumoral cells but generally presented a GI50 value higher than that of the tumor cell lines.
- Composição fenólica e atividade biológica de extratos de casca e ramos de Juniperus communis L. (zimbro comum)Publication . Xavier, Virginie; Finimundy, Tiane C.; Amaral, Joana S.; Pinela, José; Calhelha, Ricardo C.; Oliveira, Izamara; Madiavilla, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Heleno, Sandrina A.; Barros, LillianA espécie Juniperus communis L., comumente designada “zimbro” em Portugal, é uma conífera, pertencente à familia Cupressaceae e adaptada à baixa disponibilidade de nutrientes no solo. O zimbro foi usado na medicina tradicional para tratar infeções urinárias, dermatites ou como diurético, entre outros, dada a sua composição em bioativos como compostos fenólicos, terpenóides e ácidos orgânicos. Nos últimos anos, vários estudos reportam o seu potencial antimicrobiano, antioxidante, antiinflamatório, efeitos anti-diabéticos, hipocolesterolémicos e ainda a sua ação neuroprotetora, bem como capacidade antiproliferativa contra células cancerígenas e a capacidade de ativar mecanismos hepatorrenais e gastroprotetores [1,2]. No âmbito do presente trabalho, que surge no contexto do projeto BBI-JU BeonNAT, a casca do zimbro foi separada da parte aérea e posteriormente moída à parte da restante biomassa (parte aérea composta por ramos e agulhas) para comparar estas duas frações ao mesmo tempo que se pretende aproveitar todas as partes da planta, promovendo um conceito de economia circular. Depois de moídas, as amostras foram extraídas por maceração e a composição fenólica de cada fração de biomassa foi analisada por HPLC-DAD-ESI/MS, onde 17 compostos foram identificados na casca e 14 compostos na restante biomassa. Enquanto que na casca foram apenas idenditificados flavan-3-ols, na parte aérea a maior parte dos compostos são flavonoides (10 identificados), seguido de 2 flavan-3-ois e ainda 2 ácidos fenólicos, sendo o composto maioritário, nos dois casos, um dímero de procyanidina. Para avaliar a biotividade dos extratos, realizou-se a análise antimicrobiana por microdiluição, utilizando bacterias Gram-positivas e Gram-negativas, isoladas de ambiente clínico e alimentar. A atividade antioxidante foi realizada por duas metodologías distintas: inibição da peroxidação lipídica em tecidos cerebrais de porco (TBARS) e inibição da hemólise oxidativa (OxHLIA), onde o extrato da casca de zimbro se mostrou mais promissor, apresentando um EC50 mais baixo (TBARS: EC50=11.300.88 g/mL e OxHLIA: 14.9±0.5 g/mL) em comparação com o extracto da parte aérea (TBARS: EC50=25.890.62 g/mL e OxHLIA: 47±1 g/mL). Por sua vez, a citotoxicidade foi avaliada numa cultura de células não tumorais: PLP2 (cultura primária de células de fígado de porco), em que ambos os extratos apresentaram toxicidade para esta linha celular. No entanto, de acordo com a aplicação-alvo, a toxicidade apresentada pelos extratos deve ser analisada mais profundamente, verificando esta condição em modelos de toxicidade específicos para cada indústria/produto. No geral, tendo em conta a composição química e as bioatividades comprovadas, os extratos quer da casca, quer da parte aérea do zimbro são potenciais matérias-primas para o desenvolvimento de produtos com aplicação na área alimentar, farmacêutica e/ou cosmética.
- Development of bio-based ingredients from underused trees and shrub species for industrial applicationPublication . Xavier, Virginie; Heleno, Sandrina A.; Prieto Lage, Miguel A.; Amaral, Joana S.; Mediavilla Ruiz, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianAround the world, the strategies of companies and governments are increasingly converging around the concept of using biomass in industry. Besides the benefit from moving away from fossil-based raw materials, the use of natural matrices bring health properties and functionalities to the final products and is desirable from a circular economy perspective. This is leading industries like the food, cosmetic and pharmaceutic to look for bio-based ingredients to obtain these bio-based products [1]. To not compete with the current use of biomass, one strategy to obtain these natural ingredients could be the use of underutilized species cultivated in marginal lands. This research has different purposes such as the chemical characterization of selected natural matrices from Germany, Spain and Romania (Betula pendula, Calluna vulgaris, Carpinus betulus, Cistus ladanifer, Cupressus sempervirens, Cytisus scoparius, Euonymus europaeus, Hippophae rhamnoides, Juniperus communis, Populus nigra, Robinia pseudoacacia, Rosmarinus officinalis, Rubus fruticosus, Spartium junceum, Ulmus minor and Ulmus pumila) to know and improve the contents in the target compounds (triterpenoids, phenolic compounds, including flavonoids and proanthocyanidins, hydrosoluble tannins, among others). After the chemical characterization, the extraction procedures can be optimized using experimental design tools based on response surface models to obtain high yields and highly pure essential oils and polar extracts. The obtained responses will be monitored based on extraction yield and HPLC-DAD-MS analysis. The optimized extracts in the target compounds will be refined using membrane technologies in ultrafiltration processes, semi-preparative and preparative chromatography, selective precipitation and crystallization, adsorbent bed purification to obtain purified fractions or isolated compounds. To monitor the efficiency of each process and select the most adequate methodology, HPLC-DAD will be applied. From the final extracts and/or isolated compounds, a fully characterization of the bioactive potential will be done, namely through the antioxidant, antimicrobial, cytotoxic, and anti-inflammatory properties. From these bioactive screening, each developed extract and/or isolated compound will be analyzed according to different market requirements and regulatory documents to determine their applications as nutraceuticals, plant-based medicaments, feed additives and cosmetic ingredients.
- Extraction of essential oils from the residues of two shrub species aiming for their revalorization: chemical characterization and antioxidant, antimicrobial and cytotoxic activitiesPublication . Xavier, Virginie; Heleno, Sandrina A.; Prieto Lage, Miguel A.; Amaral, Joana S.; Mediavilla Ruiz, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Barros, LillianIn recent years, essential oils have been subject of research for their bioactive properties, such as antimicrobial, fungitoxic, anti-viral, anti-inflammatory and antioxidant activities. Owing to these properties they are potentially interesting for diverse industries including the food industry since one of its main problems concerns microbes and associated toxins that are responsible for food spoilage. Although the application of essential oils in the food industry may have some limitations, such as impact on the organoleptic properties and low solubility, different delivery strategies such as nanoencapsulation, active packaging and coatings are promising technologies that may overcome these issues without compromising nutritional properties in food systems [1]. In this view, increased knowledge on the composition and activity of different essential oils is needed, particularly regarding novel potential sources of essential oils such as agricultural wastes or species grown in marginal lands, on a perspective of circular economy. Therefore, in the scope of the BeonNAT project, biomass from different tree and shrub species are being screened as possible sources of essential oils and respective bioactivity evaluated. In this work, the essential oil extracted by steam distillation from the branches (>20mm) of two shrub species grown in Spain, Juniperus communis L. and Cistus ladanifer L., was characterized for their chemical composition by gas chromatography coupled with mass spectrometry (GC-MS) as well as for their antioxidant, antimicrobial, antiinflammatory and cytotoxic activities. GC-MS analysis allowed the identification of 98.1% of compounds in J. communis EO, corresponding to a total of 63 identified compounds, alfa-pinene being the major compound (35.1%), followed by limonene (15.0%), sabinene (6.7%), cis-tujopsene (8.0%), beta-myrcene (3.2%) and beta-caryophyllene (3.5%). In general, the chemical composition is in agreement with that of juniper berry EO, defined in the European Pharmacopoeia and ISO 8897 standard, except for limonene (15.0%) which was slightly higher than the defined range (Eur. Ph of 2-12% and ISO standard of 2-8%). For C. ladanifer EO, 61 compounds were identified corresponding to 92.8% of the total compounds, with viridiflorol being the main compound (24.0%), followed by alfa-pinene (19.3%), ledol (6.9%), camphene (6.7%) and bornyl acetate (5.0%), which is in good agreement with previous data [2]. Both oils showed potential against the panel of bacteria selected according to their importance in public health and foodborne diseases, highlighting the rock-rose EO that showed interesting activity against Escherichia coli, Morganella morganii, Pseudomonas aeruginosa, Enterococcus faecalis, Listeria monocytogenes and methicillin resistant Staphylococcus aureus in a concentration range of 0.039-2.5%(v/v). Regarding the antioxidant activity, both oils showed promising results, with EC50 values of 1.35 ± 0.19 mg/mL and 1.30 ± 0.07 mg/mL in the reducing power assay and 68% and 83% inhibition of oxidation according to the cellular antioxidant activity assay, for J. communis and C. ladanifer, respectively. The essential oils showed antiinflammatory (IC50 of 24±1 µg/mL and 21±2 µg/mL for juniper and rock-rose-rose, respectively) and cytotoxic activity, with the best results obtained with the rock-rose EO in the inhibition of stomach-AGS, colon-CaCo, breast-MCF-7 and lung-NCI-H460 cancer cell lines (GI50 between 47±5 µg/mL and 58±1 µg/mL). Juniper EO did not evidenced cytotoxicity in non-tumoral Vero cells at the highest tested concentration (400 µg/mL) wich can be an indicator of its safety. Overall, the results demonstrated that shrubs biomass can be a source of EO with similar composition to that reported for respective berries and leaves. The EOs showed interesting antibacterial and antioxidant activity thus being potential candidates for further studies on their safety and potential application in food systems.
- Valorização de espécies mediterrânicas subutilizadas: obtenção de óleo essencial com propriedades bioativasPublication . Xavier, Virginie; Finimundy, Tiane C.; Amaral, Joana S.; Calhelha, Ricardo C.; Vaz, Josiana A.; Pires, Tânia C.S.; Madiavilla, Irene; Esteban Pascual, Luis Saul; Ferreira, Isabel C.F.R.; Heleno, Sandrina A.; Barros, LillianAs espécies C. sempervirens L. e J. communis L. são amplamente plantadas como arbustos ornamentais em parques e jardins, enquanto C. ladanifer L. é frequentemente encontrada em áreas silvestres. No entanto, estas espécies apresentam um elevado potencial para serem cultivadas em terrenos marginais [1]. Diferentes estudos investigaram a composição química dessas espécies por GC-MS; no entanto, a maioria refere-se ao óleo esencial (OE) obtido por hidrodestilação das folhas de C. ladanifer e C. sempervirens, assim como de bagas de J. communis, sendo escassa a informação sobre outras partes das plantas [2,3]. No âmbito do projeto BBI-JU BeonNAT, o presente estudo teve como objetivo avaliar a composição química de OE extraídos por destilação a vapor da parte aérea dessas três espécies e as suas propriedades bioativas (antioxidante, antibacteriana, citotóxica e anti-inflamatória) tendo como finalidade o desenvolvimento de produtos de base biológica em diferentes indústrias. Assim, cada espécie foi colhida em duas origens diferentes para procurar diferentes quimiotipos associados a diferentes localizações geográficas [4]. Os resultados obtidos sugerem que a colheita dessas espécies em diferentes localizações geográficas interfere no rendimento do OE e respectiva composição química, sendo que o maior teor de terpenos foi observado em C. sempervirens seguido de J. communis. Com exceção de uma das origens de C. ladanifer que apresentou viridiflorol como o composto mais abundante, as três espécies apresentaram em comum o -pineno como composto majoritário. Os OEs de C. ladanifer apresentaram potencial antibacteriano, apresentando valores de concentração mínima inibitória (CMI) de 0.3 a 1.25 mg/mL. Relativamente às restantes bioatividades, o OE de C. ladanifer, revelou uma inibição da oxidação de 83%, enquanto que o OE de J. communis apresentou citotoxicidade na linhagem celular MCF-7, e os OEs C. sempervirens e C. ladanifer exibiram o maior potencial na linha celular NCI-H460. Qualquer um dos óleos apresentou actividade antiinflamatória, destacando-se o maior potential na especie de C. sempervirens (IC50: 11-14 g/mL), seguido do OE de C. ladanifer (IC50: 19-21 g/mL) e J. communis (IC50: 24-85 g/mL). As espécies avaliadas revelaram ser uma fonte viável e de baixo custo de OEs que podem ser utilizados para o desenvolvimento de produtos de base biológica em diferentes industrias.
- Yield, chemical composition and bioactivity of essential oils from common juniper (Juniperus communis L.) from different spanish originsPublication . Esteban Pascual, Luis Saul; Mediavilla, Irene; Xavier, Virginie; Amaral, Joana S.; Pires, Tânia C.S.P.; Calhelha, Ricardo C.; Lopez, Cesar; Barros, LillianEssential oils (EOs) obtained from Juniperus communis L. are frequently used in the production of bioproducts. However, there are no studies regarding industrial crops' production, allowing for better control of the quality and production of juniper EOs. To select the plant material for developing future crops of this species in northern Spain, four locations where this shrub species grows in the wild were selected and samples of both genera were collected. The EOs were obtained by steam distillation, and their chemical composition and bioactivity were evaluated. The results showed that the yield of EOs from male and female samples were within the usual reported ranges, varying between 0.24 and 0.58% (dry basis, d.b.). However, limonene content in three locations varied between 15 and 25%, which is between 100% and 200% higher than the values usually reported for other European countries. The antibacterial activity was determined by broth microdilution and showed that gram-positive bacteria were more susceptible to the tested EOs since, in general, lower MIC values were obtained compared to gram-negatives. The EOs from location 1 (L1F) and 2 (L2M) inhibited the growth of six out of the eight clinical strains tested. Samples from location 1 were particularly effective, exhibiting MBC against two gram-negative (E. coli and P. mirabilis) and one gram-positive bacteria (E. faecalis). Moreover, the majority of the EOs tested showed anti-inflammatory activity. Cytotoxic effect has been demonstrated in tumor cell lines, with the best results observed against gastric carcinoma (AGS) cells (GI(50) between 7 to 77 mu g/mL). Although generally presenting higher GI50, most samples also inhibited the growth of non-tumoral cells, particularly hepatocytes (PLP2 cells). Therefore, its use for their anti-proliferative activity should consider specific conditions to avoid damaging normal cells. Finally, the results and conclusions obtained led to the selection of the female shrubs from location 1 (L1F) as the plant material to be propagated in order to produce plants for a future juniper crop.