Betulinic Acid Nanogels: Rheological, Microstructural Characterization and Evaluation of their Anti-inflammatory Activity.

BACKGROUND: Betulinic Acid (BA) is a lipophilic compound with proven beneficial results in topical inflammation. Nanogels (NG) are carriers of bioactive compounds with properties that make them good candidates to treat skin diseases. OBJECTIVE: The objective of this study was to evaluate the anti-inflammatory activity of BA carried in NG. METHODS: NG were composed of a nanoemulsion and a crosslinking agent (Carbopol 940®) applied at three concentrations (0.5, 1, and 1.5 %) and three activation times (6, 12 and 24 h). In order to select the optimal formulation, the NG were characterized mechanically and micro-structurally followed by evaluation of the BA anti-inflammatory activity in an in vivo model of auricular edema. We determined the edema inhibition activity as percent weight. Additionally, the anti-inflammatory activity of NG was validated through histological analysis. RESULTS: The formulation with the best viscoelastic properties was the one prepared with 0.5% carbopol and 6 h of activation. Microstructural examination of this formulation showed mostly spherical structures with a mean diameter of 65 nm. From the evaluation of edema and the histological analyses, we established that the NG of BA produced 52% inhibition. In contrast, a conventional gel and free BA produced 28% and 19% inhibition, respectively. CONCLUSION: The NG of BA were found to be good vehicles to treat skin inflammation.

Lipoxygenase and Its Relationship with Ethylene During Ripening of Genetically Modified Tomato (Solanum lycopersicum).

RESEARCH BACKGROUND: TomloxB is the main isoform of lipoxygenase associated with ripening and senescence of fruits. On the other hand, ethylene, a gaseous hormone, is essential for the regulation of ripening in climacteric fruits like tomatoes. However, the relationship between TomloxB and ethylene production has not been thoroughly studied. Therefore, we aim to assess the effect of exogenous ethylene in transgenic tomatoes that contain a silenced TomloxB gene, and subsequently evaluate lipoxygenase activity, 1-aminocyclopropane-1-carboxylic acid oxidase and ethylene production; as well as to quantify the expression of the genes encoding 1-aminocyclopropane-1-carboxylic acid oxidase and TomloxB. EXPERIMENTAL APPROACH: To investigate the effect of lipoxygenase and 1-aminocyclopropane-1-carboxylic acid oxidase activity, fruits harvested at the stages of break, turning and pink were used. Tomatoes at break stage collected from transgenic and wild type plants were used to determine ethylene production and gene expression. Genetically modified and wild type tomato fruits were exposed to 100 µL/L exogenous ethylene. Lipoxygenase activity was measured spectrophotometrically. Activity of 1-aminocyclopropane-1-carboxylic acid oxidase and ethylene production were determined by gas chromatography. Oligonucleotides for differentially expressed genes: 1-aminocyclopropane-1-carboxylic acid oxidase and TomloxB were used to determine gene expression by real-time PCR. RESULTS AND CONCLUSIONS: The data showed that silencing of TomloxB caused a reduction in lipoxygenase activity and ethylene production in tomato fruits, and also reduced 1-aminocyclopropane-1-carboxylic acid oxidase activity. Hence, the addition of exogenous ethylene increased lipoxygenase activity in all treatments and 1-aminocyclopropane-1-carboxylic acid oxidase activity only in transgenic lines at break stage, consequently there was a positive regulation between TomloxB and ethylene, as increasing the amount of ethylene increased the activity of lipoxygenase. The results suggest that lipoxygenase may be a regulator of 1-aminocyclopropane-1-carboxylic acid oxidase and production of ethylene at break stage. NOVELTY AND SCIENTIFIC CONTRIBUTION: These results lead to a better understanding of the metabolic contribution of TomloxB in fruit ripening and how it is linked to the senescence-related process, which can lead to a longer shelf life of fruits. Understanding this relationship between lipoxygenase and ethylene can be useful for better post-harvest handling of tomatoes.

Curcumin Nanoemulsions Stabilized with Modified Phosphatidylcholine on Skin Carcinogenesis Protocol.

BACKGROUND: Cancer is one of the main causes of death by disease; several alternative treatments have been developed to counteract this condition. Curcumin (diferuloylmethane), extracted from the rhizome of Curcuma longa, has antioxidant, anti-inflammatory, and anti-cancer properties; however, it has low water solubility and poor intestinal absorption. Carrier systems, such as nanoemulsions, can increase the bioavailability of lipophilic bioactive compounds. OBJECTIVE: To evaluate the effect of curcumin nanoemulsions prepared with lecithin modified with medium-chain fatty acids as an emulsifier, on the expression of the Cdk4, Ccne2, Casp8 and Cldn4 genes involved in the carcinogenesis process in K14E6 transgenic mice. METHODS: The emulsifier was prepared by interesterification of medium-chain fatty acids, pure lecithin, and immobilized phospholipase-1 on Duolite A568. An Ultraturrax homogenizer and a Branson Ultrasonic processor were used for the preparation of nano-emulsions, and a Zetasizer evaluated the particle size. qRT-PCR analysis was performed to quantify the cancer-related genes expressed in the K14E6 mice. The development and evolution of skin carcinogenesis were assessed through histological analysis to compare cell morphology. RESULTS: Ca 59% of the MCFA were incorporated via esterification into the PC within 12 hours of the reaction. An emulsifier yield used to formulate the NE of 86% was achieved. Nanoemulsions with a particle size of 44 nm were obtained. The curcumin nano-emulsion group had a 91.81% decrease in the tumorigenesis index and a reduction in tumor area of 89.95% compared to the sick group. Histological analysis showed that the group administered with free curcumin developed a microinvasive squamous cell carcinoma, as opposed to the group with nanoemulsion which presented only a slight inflammation. In gene expression, only a significant difference in Cdk4 was observed in the nanoemulsion group.

A Specific Thin Layer Chromatography Method for the Identification and Separation of Medium Chain Acylglycerols.

Medium chain fatty acids (MCFAs), have gained nutritional relevance in the past few years. They are continuously used in obtaining structured lipids like medium chain acylglycerols (MCAs) for various purposes. However, because of their chemical structure pertaining carbon chain length and the presence of saturated and unsaturated fatty acids, sensitive detection techniques are required for their correct identification and separation. In the present work, a specific thin layer chromatography (TLC) method for MCAs was developed. The proposed method consisted of the use of a mixture of hexane: acetone (70:30 v/v) as mobile phase, since it proved effectiveness for the separation of compounds of interest (MCAs) as well as having the necessary sensitivity to separate different species of monoacylglycerols (MAGs), diacylglycerols (DAGs) and triacylglycerols (TAGs) of MCFAs. For observation of the compounds, a single oxidizing agent was not sufficient, thus a combination of visualization reagents was used (first a 10 % solution of sulphuric acid in methanol followed by a 10 % solution of phosphomolybdic acid in methanol) achieving the correct visualization of the desired compounds.

Increased Postharvest Life of TomLox B Silenced Mutants of Tomato (Solanum lycopersicum) Var. TA234.

A healthy lifestyle includes fruits and vegetables consumption. Tomato is one of the most consumed vegetables, although it is susceptible to physical damage through postharvest handling, thus leading to important losses. Softening is an important variable during tomato ripening; excessive softening is undesirable and leads to postharvest losses. TomloxB plays an important role in ripening, mainly in the loss of cellular integrity caused by fatty acids released from the lipid matrix of membranes that initiate oxidative deterioration, which is in turn carried into senescence. In order to increase postharvest life, we produced transgenic tomato plants via Rhizobium radiobacter with tomato lipoxygenase B (TomloxB) antisense constructs under control of the cauliflower mosaic virus (CaMV) 35S promoter. Lipoxygenase activity and firmness were measured in tomato fruit and the fatty acids profile was determined. Transgenic fruits were maintained for 40 days at room temperature in optimal conditions, whereas wild type fruits remained in similar conditions for only six days. Firmness in pink and red stages was significantly lower in wild type fruits than in two transgenic lines. Linolenic acid was the most important fatty acid consumed by lipoxygenase in both turning and pink stages of ripening. Lipoxygenase activity was smaller in transformed fruits in comparison with the wild type. These results suggest that silencing the TomloxB gene promoted significant changes in the physiology of transformed tomatoes, being the increase in postharvest life the most important.