Thermodynamic Behavior of (2-Propanol + 1,8-Cineole) Mixtures: Isothermal Vapor-Liquid Equilibria, Densities, Enthalpies of Mixing, and Modeling.

Vapor pressures and other thermodynamic properties of liquids, such as density and enthalpy of mixtures, are the key parameters in chemical engineering for designing new process units, and are also essential for understanding the physical chemistry, macroscopic and molecular behavior of fluid systems. In this work, vapor pressures between 278.15 and 323.15 K, densities and enthalpies of mixtures between 288.15 and 318.15 K for the binary mixture (2-propanol + 1,8-cineole) have been measured. From the vapor pressure data, activity coefficients and excess Gibbs energies were calculated via the Barker's method and the Wilson equation. Excess molar volumes and excess molar enthalpies were also obtained from the density and calorimetric measurements. Thermodynamic consistency test between excess molar Gibbs energies and excess molar enthalpies has been carried out using the Gibbs-Helmholtz equation. Robinson-Mathias, and Peng-Robinson-Stryjek-Vera together with volume translation of Peneloux equations of state (EoS) are considered, as well as the statistical associating fluid theory that offers a molecular vision quite suitable for systems having highly non-spherical or associated molecules. Of these three models, the first two fit the experimental vapor pressure results quite adequately; in contrast, only the last one approaches the volumetric behavior of the system. A brief comparison of the thermodynamic excess molar functions for binary mixtures of short-chain alcohol + 1,8-cineole (cyclic ether), or +di-n-propylether (lineal ether) is also included.

Supercritical Antisolvent Fractionation of Antioxidant Compounds from Salvia officinalis.

The increasing interest towards greener antioxidants obtained via natural sources and more sustainable processes encourages the development of new theoretical and experimental methods in the field of those compounds. Two advanced separation methods using supercritical CO2 are applied to obtain valuable antioxidants from Salvia officinalis, and a first approximation to a QSAR model relating molecular structure with antioxidant activity is explored in order to be used, in the future, as a guide for the preselection of compounds of interest in these processes. Separation experiments through antisolvent fractionation with supercritical CO2 were designed using a Response Surface Methodology to study the effect of pressure and CO2 flow rate on both mass yields and capability to obtain fractions enriched in three antioxidant compounds: chlorogenic acid, caffeic acid and rosmarinic acid which were tracked using HPLC PDA. Rosmarinic acid was completely retained in the precipitation vessel while chlorogenic and caffeic acids, though distributed between the two separated fractions, had a major presence in the precipitation vessel too. The conditions predicted for an optimal overall yield and enrichment were 148 bar and 10 g/min. Although a training dataset including much more compounds than those now considered can be recommended, descriptors calculated from the σ-profiles provided by COSMO-RS model seem to be adequate for estimating the antioxidant activity of pure compounds through QSAR.

Impact of citronellol on river and soil environments using non-target model organisms and natural populations.

Citronellol is an acyclic monoterpenoid with a wide range of pharmacological activities (antibacterial, antifungal, anti-lice, repellent, lipolytic, anti-allergic, anti-inflammatory, antispasmodic, antidiabetic, anti-cholesterol, among other) and potential to replace synthetic products. However, the impact of citronellol on the environment remains unknown. We analysed, for the first time, the environmental impact of citronellol on river and soil environments using non-target model organisms and natural populations. The acute toxicity of citronellol on the aquatic invertebrate Daphnia magna, the plant Allium cepa L and the earthworm Eisenia fetida was quantified. The effect of citronellol in a river ecosystem was analysed using river periphyton communities taxonomically characterised and a river microbial community characterised through 16 S rRNA gene sequencing. Finally, a microbial community from natural soil was used to monitor the effect of citronellol on the soil ecosystem. The results showed that E. fetida was most sensitive to citronellol (LC50 = 12.34 mg/L), followed by D. magna (LC50 = 14.11 mg/L). Citronellol affected the photosynthesis of the fluvial periphyton (LC50 = 94.10 mg/L) and was phytotoxic for A. cepa. Furthermore, citronellol modified the growth and metabolism of both fluvial (LC50 = 0.19% v/v) and edaphic (LC50 = 5.07% v/v) bacterial populations. The metabolism of the microorganisms in the soil and water exposed to citronellol decreased with respect to the control, especially their ability to metabolise carbohydrates. Our results show that citronellol has a negative impact on the environment. Although acute effects cannot be expected, it is necessary to quantify the environmental levels as well as the long-term and persistent effects of this monoterpene.

Concentration of Antioxidant Compounds from Calendula officinalis through Sustainable Supercritical Technologies, and Computational Study of Their Permeability in Skin for Cosmetic Use.

The growing interest in the cosmetic industry in using compounds of natural and sustainable origin that are safe for humans is encouraging the development of processes that can satisfy these needs. Chlorogenic acid (CHA), caffeic acid (CAF) and ferulic acid (FA) are three compounds widely used within the cosmetic industry due to their functionalities as antioxidants, collagen modifiers or even as radiation protectors. In this work, two advanced separation techniques with supercritical CO2 are used to obtain these three compounds from Calendula officinalis, and these are then evaluated using a computational skin permeability model. This model is encompassed by the COSMO-RS model, the calculations of which make it possible to study the behaviour of the compounds in the epidermis. The results show that both CAF and FA are retained in the stratum corneum, while CHA manages to penetrate to the stratum spinosum. These compounds were concentrated by antisolvent fractionation with super-critical CO2 using a Response Surface Methodology to study the effect of pressure and CO2 flow rate. CHA, CAF and FA were completely retained in the precipitation vessel, with concentrations between 40% and 70% greater than in the original extract. The conditions predicted that the optimal overall yield and enrichment achieved would be 153 bar and 42 g/min.

Effects of the insecticide fipronil in freshwater model organisms and microbial and periphyton communities.

Fipronil is a broad-spectrum insecticide whose release in the environment damages many non-target organisms. This study evaluated the toxicity of fipronil at two biological levels using in vivo conditions and environmentally relevant concentrations: the first based on two model organisms (aquatic invertebrate Daphnia magna and the unicellular freshwater alga Chlamydomonas reinhardtii) and a second based on three natural communities (river periphyton and freshwater and soil microbial communities). The physicochemical properties of fipronil make it apparently unstable in the environment, so its behaviour was followed with high performance liquid chromatography (HPLC) under the different test conditions. The most sensitive organism to fipronil was D. magna, with median lethal dose (LC50) values from 0.07 to 0.38 mg/L (immobilisation test). Toxicity was not affected by the media used (MOPS or river water), but it increased with temperature. Fipronil produced effects on the photosynthetic activity of C. reinhardtii at 20 °C in MOPS (EC50 = 2.44 mg/L). The freshwater periphyton presented higher sensitivity to fipronil (photosynthetic yield EC50 of 0.74 mg/L) in MOPS and there was a time-dependent effect (toxicity increased with time). Toxicity was less evident when periphyton and C. reinhardtii tests were performed in river water, where the solubility of fipronil is poor. Finally, the assessment of the metabolic profiles using Biolog EcoPlates showed that bacteria communities were minimally affected by fipronil. The genetic identification of these communities based on 16S rRNA gene sequencing revealed that many of the taxa are specialists in degrading high molecular weight compounds, including pesticides. This work allows us to better understand the impact of fipronil on the environment at different levels of the food chain and in different environmental conditions, a necessary point given its presence in the environment and the complex behaviour of this compound.

Supercritical Carbon Dioxide Antisolvent Fractionation for the Sustainable Concentration of Lavandula luisieri (Rozeira) Riv.- Mart Antimicrobial and Antioxidant Compounds and Comparison with Its Conventional Extracts.

Lavandula stoechas subsp. luisieri is a Spanish subspecies from the Lamiaceae family. Its essential oil has been traditionally used for several medical applications though little is known about other extracts. Similar to many other studies aiming to obtain traditional plant extracts to be used in different applications, this work evaluated the antioxidant and antimicrobial activities of Lavandula luisieri extracts and the correlation with their composition. Traditional hydrodistillation and ethanolic maceration were used to obtain the essential oil and the maceration extract, respectively. A green and sustainable methodology was applied to the maceration extract that was under a Supercritical Antisolvent Fractionation process to obtain a fine solid enriched in rosmarinic acid and the terpenes oleanolic and ursolic acids. Antimicrobial activities of all extracts and pure identified compounds (rosmarinic and ursolic acids) were evaluated against five bacterial strains; Listeria monocytogenes, Enterococcus faecium, Staphylococcus aureus, Salmonella Typhimurium and Escherichia coli and were compared with the pure compounds identified, rosmarinic and ursolic acids. All strains were sensitive against L. luisieri essential oil. The solid product obtained from the supercritical process was concentrated in the identified actives compared to the maceration extract, which resulted in higher antimicrobial and DPPH scavenging activities. The supercritical sustainable process provided L. luisieri compounds, with retention of their antimicrobial and antioxidant activities, in a powder exemptof organic solvents with potential application in the clinical, food or cosmetic fields.

Impact of Artemisia absinthium hydrolate extracts with nematicidal activity on non-target soil organisms of different trophic levels.

Natural pesticides are considered a good alternative to synthetic pesticides to reduce environmental impacts. However, biopesticides may have unknown effects on the environment, and can affect non-target organisms. In this study, the ecotoxicological effects of an aqueous extract (hydrolate) from Spanish populations of Artemisia absinthium (var. Candial) showing a promising biopesticide activity, were evaluated on non-target soil organisms from different trophic levels (natural microbial communities characterized through 16S rRNA gene sequencing, the earthworm Eisenia fetida and the plant Allium cepa). The hydrolate usually was considered as a by-product of the distillation to obtain essential oils. However, recently has been found to have nematicide properties. The hydrolate caused acute toxicity at values of LC50 of 3.87% v/v for A. cepa and 0.07 mL/g for E. fetida. All the concentrations except for the most diluted (1% v/v) reduced the bacterial physiological activity compared to controls (LC50 = 25.72% v/v after 24 h of exposure). The hydrolate also slightly altered the ability of the microbial community to degrade carbon substrates. These results indicate that the hydrolate from A. absinthium may affect the survival and metabolic abilities of key soil organisms.

Ecotoxicity of a new biopesticide produced by Lavandula luisieri on non-target soil organisms from different trophic levels.

Plant-based biopesticides have become an eco-friendly alternative to synthetic pesticides by reducing the undesired environmental impacts and side-effects on human health. However, their effects on the environment and especially on non-target organisms have been little studied. This study analyses the ecotoxicological effects of the extract of Lavandula luisieri on soil non-target organisms from different trophic levels: the earthworm Eisenia fetida, the plant Allium cepa and a natural-soil microbial community whose taxonomy was analysed through 16S rRNA gene sequencing. The extract tested is the hydrolate -product from a semi industrial steam distillation process- of a Spanish pre-domesticated variety of L. luisieri. This hydrolate has been recently shown to have bionematicide activity against the root-knot nematode Meloidogyne javanica. A previous study showed that the main components of the hydrolate are camphor and 2,3,4,4-Tetramethyl-5-methylidenecyclopent-2-en-1-one. Hydrolate caused acute toxicity (LC50 2.2% v/v) on A. cepa, while only a slight toxicity on E. fetida (LC50 > 0.4 mL/g). All the concentrations tested (from 1 to 100% v/v) caused a significant decrease in bacterial growth (LC50 9.8% v/v after 120 h of exposure). The physiological diversity of the community was also significantly altered, except in the case of the lowest concentration of hydrolate (1% v/v). The ability of soil microbial communities to use a variety of carbon sources increased for all substrates at the highest concentrations. These results show that both the plants and bacterial communities of the soil can be affected by the application of biopesticides based on these hydrolates, which highlights the need for a more detailed risk assessment during the development of plant-based products.

Ecotoxicity of a novel biopesticide from Artemisia absinthium on non-target aquatic organisms.

Biopesticides are increasingly being used to replace synthetic pesticides for pest control. This change raises concern for its environmental impacts, especially on non-target organisms. In this study, the ecotoxicological effects of a potential nematicide from Spanish populations of Artemisia absinthium (var. Candial) were evaluated on freshwater and aquatic non-target organisms. The study focused on the aqueous extract (hydrolate), the principal component of which ((-) -(Z) -2,6-dimethylocta-5,7-diene-2,3-diol) is responsible for its nematicidal effect. Until now, the hydrolate has been considered a byproduct of the process used to obtain essential oils, and there are no studies on its ecotoxicity from any plant with biopesticide properties. Our results indicated that A. absinthium hydrolate caused acute toxicity for non-target organisms at dilutions as low as 0.2%. The sensitivity of the organisms, from the most to the least sensitive, was: Daphnia magna (LC50 = 0,236%) > Vibrio fisheri (LC50 = 1,85%) > Chlamydomonas reinhardtii (LC50 = 16,49). Moreover, the A. absinthium organic extract was highly toxic to D. magna (LC50 = 0,093 mg/L). A. absinthium hydrolate toxicity was also tested on a natural river microbial community. Bacterial growth was not affected; the physiology of the community was only slightly modified, namely through an increased ability to degrade different substrates, mainly carbohydrates. This study provides for the first time an exhaustive assessment of the environmental exposure of a plant-derived biopesticide and shows that these products may cause a broad range of toxicity on non-target aquatic organisms.

Acute toxicological effects on the earthworm Eisenia fetida of 18 common pharmaceuticals in artificial soil.

Following soil applications of recycled water and biosolids, pharmaceutical residues can eventually enter the terrestrial environment. In vitro and in vivo assays have largely focused on the acute ecotoxicity of these compounds in aquatic systems. However, studies on the ecotoxicological effects of pharmaceuticals in soil biota are especially scarce. The aim of this study was to investigate the acute toxicity of 18 pharmaceuticals (4 NSAIDs, 5 blood lipid-lowering agents, 6 ß-blockers and 3 antibiotics) that are usually found in the environment by using an Eisenia fetida bioassay. In addition, the presence of these pharmaceuticals in artificial soil was verified at the end of the test. Our results indicate that seven of the studied drugs cause acute adverse effects in E. fetida, in particular, the NSAIDs and the blood lipid-lowering agents. Ibuprofen (LC50=64.80 mg/kg) caused the highest acute toxicity for all tested compounds, followed by diclofenac (LC50=90.49 mg/kg) and simvastatin (LC50=92.70 mg/kg). Other tested pharmaceuticals from NSAIDs and blood lipid-lowering families have toxicity effects, from a LC50=140.87 mg/kg for gemfibrozil to 795.07 mg/kg for lovastatin. Atorvastatin, bezafibrate, ß-blockers and antibiotics showed no detectable lethality in E. fetida. The four NSAIDs showed evidence of modification of their original chemical structure after 14 days so the detected toxicity may be due to the original product as well as their degradation products. The three blood lipid-lowering agents seem to be more stable in soil. From an environmental perspective, the lethal concentrations of the tested drugs are much greater than those reported in wastewater and biosolids, therefore acute toxic effects may be improbable. However, little is known about the accumulation of these substances in soils after regular applications, so accumulative and chronic effects cannot be excluded. Moreover, more studies are needed to determine the role of the degradation products of these pharmaceuticals on terrestrial toxicity.

A new generation of cysteine derivatives with three active antioxidant centers: improving reactivity and stability.

The development of new antioxidants with enhanced activity constitutes a very active research field as it can contribute to the improvement of human health. Although the antioxidant activity occurs through different mechanisms, usually most of the antioxidant molecules present a unique active center which is able to react following a specific way. To overcome this weakness and in the belief that the coupling of different antioxidant groups is a good strategy to obtain multipotent antioxidants, the effect of introducing different N-protective groups on the cysteine core is evaluated by using DFT. As a result, in this work we present a multicenter antioxidant, N-(9-fluorenylmethyloxycarbonyl)cysteine methyl ester 8, able to fight efficiently through different mechanisms against free radicals independently of their nature. This antioxidant appears to be the first one of a promising new class of multipotent antioxidants with three operative centers: C(α) that is a good hydrogen donor, the Fmoc group that is a good electron donor and the all-around thiol group. Besides, its neutral radical shows a high stability due to the captodative effect in such a way that the subsequent toxic effects would be avoided. Then, its experimental radical-trapping antioxidant activity postulates compound 8 as a prototype of antioxidants more versatile and efficient than N-acetylcysteine, ascorbic acid or Trolox.

Intramolecular hydrogen-bonding activation in cysteines: a new effective radical scavenger.

The challenge of developing organic molecules with improved antioxidant activities for a competitive marketplace requires, given the great amount of possibilities, much laboratory work. Nowadays, the ability of methodologies based on quantum chemistry to determine the influence of different modifications on a molecule core provides a powerful tool for selecting the most useful derivatives to be synthesized. Here, we report the results of the assessment of antioxidant activity for quaternary amino acids, specifically for cysteine derivatives. The effect of introducing different substituents on the cysteine core is evaluated by using DFT to obtain an adequate structure-antioxidant activity relationship. This theoretical study shows a small panel of targets among which (R)-N-acetyl-2-methylcysteine methyl ester 15 exhibits special features and relevant antioxidant activity. The conformational (1)H NMR study of this synthesized compound indicates the existence of an intramolecular C7 member ring involving S-HO[double bond, length as m-dash]C substructure, which is reported for the first time in the literature for this amino acid unit. This unusual conformation seems to be the reason for the high antioxidant capacity experimentally found for this compound.

Extraction of volatile oil from aromatic plants with supercritical carbon dioxide: experiments and modeling.

An overview of the studies carried out in our laboratories on supercritical fluid extraction (SFE) of volatile oils from seven aromatic plants: pennyroyal (Mentha pulegium L.), fennel seeds (Foeniculum vulgare Mill.), coriander (Coriandrum sativum L.), savory (Satureja fruticosa Béguinot), winter savory (Satureja montana L.), cotton lavender (Santolina chamaecyparisus) and thyme (Thymus vulgaris), is presented. A flow apparatus with a 1 L extractor and two 0.27 L separators was built to perform studies at temperatures ranging from 298 to 353 K and pressures up to 30.0 MPa. The best compromise between yield and composition compared with hydrodistillation (HD) was achieved selecting the optimum experimental conditions of extraction and fractionation. The major differences between HD and SFE oils is the presence of a small percentage of cuticular waxes and the relative amount of thymoquinone, an oxygenated monoterpene with important biological properties, which is present in the oils from thyme and winter savory. On the other hand, the modeling of our data on supercritical extraction of volatile oil from pennyroyal is discussed using Sovová's models. These models have been applied successfully to the other volatile oil extractions. Furthermore, other experimental studies involving supercritical CO(2) carried out in our laboratories are also mentioned.

Quantum-chemical predictions of redox potentials of carbamates in methanol.

Redox potentials for two stepwise anodic oxidations of a series of carbamates in methanolic solution have been calculated using ab initio and DFT quantum mechanical methods. Hartree-Fock method and Density Functional Theory at the B3LYP level, together with 6-31G(d), 6-31G(d,p) and 6-311++G(2df,2p) basis sets have been used for the calculation. The Polarizable Continuum Model (PCM) is used to describe the solute-solvent interactions of carbamates, and those of their radical-cations and cations. Linear relationships were observed between the theoretically predicted redox potential values and the corresponding anodic peak potentials obtained by cyclic voltammetry or the corresponding calculated energies of the Highest Occupied Molecular Orbital (HOMO) of these carbamates.

Composition and antioxidant activity of Thymus vulgaris volatiles: comparison between supercritical fluid extraction and hydrodistillation.

Supercritical fluid extraction (SFE) of the volatile oil from Thymus vulgaris L. aerial flowering parts was performed under different conditions of pressure, temperature, mean particle size and CO(2) flow rate and the correspondent yield and composition were compared with those of the essential oil isolated by hydrodistillation (HD). Both the oils were analyzed by GC and GC-MS and 52 components were identified. The main volatile components obtained were p-cymene (10.0-42.6% for SFE and 28.9-34.8% for HD), gamma-terpinene (0.8-6.9% for SFE and 5.1-7.0% for HD), linalool (2.3-5.3% for SFE and 2.8-3.1% for HD), thymol (19.5-40.8% for SFE and 35.4-41.6% for HD), and carvacrol (1.4-3.1% for SFE and 2.6-3.1% for HD). The main difference was found to be the relative percentage of thymoquinone (not found in the essential oil) and carvacryl methyl ether (1.0-1.2% for HD versus t-0.4 for SFE) which can explain the higher antioxidant activity, assessed by Rancimat test, of the SFE volatiles when compared with HD. Thymoquinone is considered a strong antioxidant compound.

Supercritical fluid extraction of the volatile oil from Santolina chamaecyparissus.

Supercritical fluid extraction (SFE) of the volatile oil from Santolina chamaecyparissus L. flower heads was performed under different conditions of pressure, temperature, mean particle size and CO(2) flow rate. This oil was compared with the essential oil isolated by hydrodistillation (HD). The SFE volatile and essential oils were analysed by GC and GC-MS. The range of the main volatile components obtained with HD and SFE were, respectively: 1,8-cineole (25-30% and 7-48%), camphor (7-9% and 8-14%), borneol (7-8% and 2-11%), terpinen-4-ol (6-7% and 1-4%), terpinolene (1-4% and 1-7%) and isobornyl acetate (1-2% and 1-11%). The chemical composition of the extracts was greatly influenced by the conditions of pressure and temperature used. In fact, it was possible to enrich the sesquiterpene fraction by increasing the pressure from 8 to 9 MPa, while changing the temperature from 50 to 40 degrees C at 9 MPa enriched the volatiles in n-alkanes [corrected].

Enrichment of the thymoquinone content in volatile oil from Satureja montana using supercritical fluid extraction.

Supercritical fluid extraction (SFE) of the volatile oil from Satureja montana L. was performed under different conditions of pressure (90 and 100 bar), temperature (40 and 50 degrees C), mean particle sizes (0.4, 0.6 and 0.8 mm) and CO(2) flow rate (0.8, 1.1 and 1.3 kg/h) to understand the influence of these parameters on the composition and yield of this oil. The results were compared with those obtained for the essential oil isolated by hydrodistillation (HD). The volatile and the essential oil were analysed by GC and GC-MS. The main compounds are carvacrol (52.2-62.0% for HD vs. 41.7-64.5% for SFE), thymol (8.6-11.0% for HD vs. 6.0-11.3% for SFE), p-cymene (6.9-12.8% for HD vs. 6.0-17.8% for SFE), gamma-terpinene (6.4-9.4% for HD vs. 2.3-6.0% for SFE) and beta-bisabolene (2.0-2.7% for HD vs. 2.2-3.5% for SFE). The major difference between SFE and HD was the relative amount of thymoquinone, an oxygenated monoterpene with important biological activities, which can be ten-fold higher in volatile oil (1.6-3.0 for SFE vs. 0.2% for HD). The morphology of the glandular trichomes of S. montana and the effect of the grinding process on them was also evaluated by SEM.