Unveiling the formation capacity of multicomponent oleogels: Performance of lecithin interacting with monostearate derivatives.

Oleogels have been explored as fat substitutes due to their healthier composition compared to trans and saturated fats, also presenting interesting technological perspectives. The aim of this study was to investigate the compositional perspective of multicomponent oleogels. Structuring ability of lecithin (LEC) (20 or 90 wt% of phosphatidylcholine - PC) combined with glycerol monostearate (GMS), sorbitan monostearate (SMS) or sucrose monostearate (SAC) in sunflower oil was evaluated from oleogels properties. The thermal and rheological properties, microstructure and stability of the oleogels were affected by the difference in the chemical composition of LEC and the ratio between LEC and different surfactants. Interestingly, low-phosphatidylcholine LEC (L20) performed better, although systems formed with reduced amounts of LEC tended to be softer (LEC-GMS) and present high oil holding capacity (LEC-SMS). The mixtures of LEC and monostearate-based surfactants showed different behaviors, depending on the surfactant polar head. In LEC-GMS systems, LEC hindered the self-assembly of GMS in sunflower oil, compromising mechanical properties and increasing oil release. When combined with SMS, LEC acted as a crystal habit modifier of SMS, forming a more homogeneous microstructure and producing stronger oleogels with greater oil binding capacity. However, above the threshold concentration, LEC prevented SMS self-assembly, resulting in a weaker gel. A positive interaction was found in LEC-SAC formulations in specific ratios, since SAC cannot act as a single oleogelator. Results show the impact of solubility balance played by LEC and fatty-acid derivatives surfactant when combined and used as oleogelators. This knowledge can contribute to a rational perspective in the preparation and modulation of the properties of edible oleogels.

The Metabolism of 2-arachidonoylglycerol in Rod Outer Segments Is Modulated by Proteins Involved in the Phototransduction Process.

We previously reported that 2-arachidonoylglycerol (2-AG) synthesis by diacylglycerol lipase (DAGL) and lysophosphatidate phosphohydrolase (LPAP) and hydrolysis by monoacylglycerol lipase (MAGL) in rod outer segments (ROS) from bovine retina were differently modified by light applied to the retina. Based on these findings, the aim of the present research was to evaluate whether 2-AG metabolism could be modulated by proteins involved in the visual process. To this end, ROS kept in darkness (DROS) or obtained in darkness and then subjected to light (BROS) were treated with GTPγS and GDPßS, or with low and moderate ionic strength buffers for detaching soluble and peripheral proteins, or soluble proteins, respectively. Only DAGL activity was stimulated by the application of light to the ROS. GTPγS-stimulated DAGL activity in DROS reached similar values to that observed in BROS. The studies using different ionic strength show that (1) the highest decrease in DROS DAGL activity was observed when both phosphodiesterase (PDE) and transducin α (Tα) are totally membrane-associated; (2) the decrease in BROS DAGL activity does not depend on PDE association to membrane, and that (3) MAGL activity decreases, both in DROS and BROS, when PDE is not associated to the membrane. Our results indicate that the bioavailability of 2-AG under light conditions is favored by G protein-stimulated increase in DAGL activity and hindered principally by Tα/PDE association with the ROS membrane, which decreases DAGL activity.

Butyric acid glycerides as substitutes for antibiotics as growth enhancers in the diet of nursery piglets.

This study aimed to determine whether the addition of butyric acid glycerides as substitutes to conventional growth promoters can provide adequate zootechnical performance and intestinal health in healthy piglets in the nursery phase. We used 90 male piglets (average weight of 6.5 kg) subdivided into five treatments with six replicates per treatment. The treatments had the same basal diet: NC-negative control (without growth promoter), PC-positive control (with gentamicin, oral), PSB-protected sodium butyrate, FSB-free sodium butyrate, and TRI-tributyrin. In these animals, zootechnical performance was evaluated on days 1, 10, 20 and 39, microbiological analysis on days 14 and 39, hematocrit, blood biochemistry and intestinal histology, intestinal oxidation and antioxidation on day 39. The average daily weight gain was higher in the TRI group on days 21 to 39 in the nursery (P = 0.03), with more significant weight gain from 1 to 39 days (P = 0.05). There were higher leukocyte counts in the PC group than in the TRI group and higher lymphocyte counts in the PC treatment than in the NC or TRI groups. Escherichia coli counts were lower in the PC, followed by the PSB and TRI groups on day 39 (P = 0.01). Lower crypt depths were found in the TRI and FSB groups, followed by PC, than in the NC group (P = 0.01). Higher values for crypt villosity ratio were found in the FSB and TRI groups than in the NC group (P = 0.05). Lower lipid peroxidation was found in analyzes of serum oxidative status (LPO: P = 0.01), associated with greater activities of superoxide dismutase - SOD (P = 0.08), glutathione S-transferase - GST (P = 0.09) in PSB and TRI groups than in the NC group. In conclusion, the use of butyric acid in the form of tributyrin can be used as growth enhancers in piglets in the nursery phase.

Drug-excipient compatibility studies in formulation development: A case study with benznidazole and monoglycerides.

Monoglycerides (MGs) such as glycerol monolaurate (GML) and glycerol monostearate (GMS) have been used as excipients in oral formulations because of their emulsifying effect as well as their ability to inhibit the precipitation and intestinal efflux of drugs. Excipient-drug compatibility studies, however, have been underexplored. In this study, benznidazole (BNZ) was selected as a drug model due to the difficulty in improving its solubility and because of the potential impact on public health (it is the only drug currently used to treat Chagas disease). The effect of different processing conditions (maceration, ball milling, and melting) on the physical-chemistry properties of BNZ/MGs mixtures was investigated to guide the rational development of new solid formulations. GML was more effective in improving the solubility of BNZ, which could be due to its more malleable structure, less hydrophobic nature, and greater interaction with BNZ. The formation of hydrogen bonds between the imidazole group of BNZ and the polar region of GML was confirmed by spectroscopy analyses (IR, 1H NMR). The higher the monoglyceride content in the mixture, the higher the BNZ solubility. Regardless of the method of processing the mixture, the drug was found to be crystalline. Polarized light microscopy analysis showed the presence of spherulites. Overall, these findings suggest that preparation methods of BNZ:MGs formulations that involve thermal or/and mechanical treatment have a low impact on the solid properties of the material, and this allows for the production of formulations with reproducible performance.

Effect of Structured Phenolic Lipids with EPA/DHA and Gallic Acid against Metabolic-Associated Fatty Liver Disease (MAFLD) in Mice.

Obesity is the leading risk factor for developing metabolic (dysfunction)-associated fatty liver disease (MAFLD). The food industry has an essential role in searching for new strategies to improve primary food sources to revert some of the metabolic alterations induced by obesity. There is consistent evidence that long-chain polyunsaturated fatty acids (n-3 LCPUFA) belonging to the n-3 series, i.e., eicosapentaenoic (20:5n-3, EPA) and docosahexaenoic (22:6n-3, DHA) acids, could revert some alterations associated with obesity-induced metabolic diseases. A relevant tool is the synthesis of structured acylglycerols (sAG), which include EPA or DHA at the sn-2 position. On the other hand, it has been reported that a crucial role of antioxidants is the reversion of MAFLD. In this work, we studied the effects of new molecules incorporating gallic acid (GA) into EPA/DHA-rich structured lipids. Mice were fed with a high-fat diet (60%) for three months and were then divided into five groups for supplementation with sAG and sAG structured with gallic acid (structured phenolic acylglycerols, sPAG). sPAG synthesis was optimized using a 2²-screening factorial design based on the response surface methodology (RSM). Our results show that treatment of sPAG was effective in decreasing visceral fat, fasting glycemia, fasting insulin, suggesting that this new molecule has a potential use in the reversal of MAFLD-associated alterations.

Acute and Chronic Toxicity of Neem Oil to the Endoparasitoid Palmistichus elaeisis (Hymenoptera: Eulophidae).

Palmistichus elaeisis Delvare and LaSalle 1993 (Hymenoptera: Eulophidae) and neem oil are two control alternatives for the integrated management of defoliating lepidopterans. The aim of this study was to evaluate the acute and chronic toxicity of neem oil compared to the synthetic insecticide deltamethrin, on the endoparasitoid P. elaeisis, in generations F0, F1, F2, and F3. Females of P. elaeisis were exposed to neem solutions at concentrations of 0, 1.87, 3.75, 7.50, 15.00, 30.00, and 60.00 mg ml-1, to determine the dose-response relationship and estimate the neem LC50. The sublethal effects on the parasitoid P. elaeisis in generations F0 to F3 were evaluated with these same concentrations of neem and 0.033 mg ml-1 of deltamethrin. The neem LC50 was estimated at 3.92 mg ml-1. The LC50 for P. elaeisis is 3.83 times lower than that recommended by the neem manufacturer for pest control, demonstrating high acute toxicity to this natural enemy. The chronic toxicity of both the commercial dose and those below it to P. elaeisis caused low sublethal effects. The correct concentration of neem oil in pest control is important, and its use should be performed with caution in integrated pest management programs using the endoparasitoid P. elaeisis to avoid causing interference between the two pest control techniques.

Butyric acid glycerides in the diet of broilers to replace conventional growth promoters: effects on performance, metabolism, and intestinal health.

This study aimed to determine whether butyric acid glycerides can replace conventional growth promoters, favour intestinal health, and improve performance. A total of 420 birds were used, divided into four groups with seven repetitions per group (n = 15), as follows: NC, negative control (no promoter); PC, positive control (basal diet + enramycin + salinomycin); MDT-BUT, a diet supplemented with mono-, di-, and triglycerides of butyric acid; TRI-BUT, a diet supplemented with tributyrin of butyric acid glycerides. Productive performance was measured on days 1, 21, 35, and 42. Excreta were collected for counting Escherichia coli and coliforms on days 21 and 42. Blood samples were collected at 42 days of age to analyse oxidant/antioxidant status, and the intestine was removed for intestinal morphometry. From 1 to 42 days, there was greater body weight, weight gain, and feed conversion in the PC, MDT-BUT, and TRI-BUT groups than in the NC group; the production efficiency index was 21.10% higher in all groups than in the NC group (p = 0.001). At 21 days, there were lower E. coli counts of 86.8% in the TRI-BUT and 99.7% in PC groups than in the NC and MDT-BUT groups (p < 0.001), while at 42 days, lower counts were found in the PC, MDT-BUT, and TRI-BUT groups than the NC group (p < 0.001). There were lower total protein and globulin levels in the MDT-BUT and TRI-BUT groups than in the NC group (p = 0.001). Cholesterol levels were lower in the TRI-BUT group, followed by MDT-BUT and PC groups, than in the NC group (p = 0.001), while lower triglyceride levels were found in the TRI-BUT group than in the NC and PC groups (p = 0.001). There were lower levels of lipid peroxidation and reactive oxygen species in the TRI-BUT group, followed by the PC group than the NC group (p < 0.001); on the other hand, there were higher protein thiol levels in the TRI-BUT group than the NC group (p = 0.041). The villus:crypt ratio increase was 79.4% in the TRI-BUT group, followed by the 45.1% PC and 19.8% MDT-BUT groups than the NC (p < 0.001). These findings suggest that adding butyric acid confers antimicrobial and antioxidant activity and improves birds' production efficiency, intestinal health, and metabolism. Butyric acid glycerides are an effective alternative to conventional growth promoters.

Anti-aversive effect of 2-arachidonoylglycerol in the dorsolateral periaqueductal gray of male rats in contextual fear conditioning and Vogel tests.

The endocannabinoid system modulates the stress coping strategies in the dorsolateral periaqueductal grey (dlPAG). The most relevant endocannabinoids, anandamide and 2-arachidonoylglycerol (2-AG) exert inhibitory control over defensive reactions mediated by the dlPAG. However, the protective role of anandamide is limited by its lack of effect in higher concentrations. Thus, the 2-AG emerges as a complementary target for developing new anxiolytic compounds. Nevertheless, the role of 2-AG on stress responsivity may vary according to the nature of the stimulus. In this study, we verified whether the dlPAG injection of 2-AG or inhibitors of its hydrolysis induce anxiolytic-like effects in male Wistar rats exposed to behavioral models in which physical stress (mild electric shock) is a critical component, namely the contextual fear conditioning test (CFC) and the Vogel conflict test (VCT). We also investigated the contribution of cannabinoid receptor type 1 (CB1) and type 2 (CB2) in such effects. The facilitation of 2-AG signaling in the dlPAG reduced contextual fear expression and exhibited an anxiolytic-like effect in the VCT in a mechanism dependent on activation of CB1 and CB2. However, the VCT required a higher dose than CFC. Further, the monoacylglycerol inhibitors, which inhibit the hydrolysis of 2-AG, were effective only in the CFC. In conclusion, we confirmed the anti-aversive properties of 2-AG in the dlPAG through CB1 and CB2 mechanisms. However, these effects could vary according to the type of stressor and the anxiety model employed.

Use of blend based on an emulsifier, monolaurin, and glycerides of butyric acid in the diet of broilers: impacts on intestinal health, performance, and meat.

The objective of this study was to evaluate whether the addition of a blend based on α-monolaurin mono-, di- and triglycerides of butyric acid, and lysolecithin on the performance even on diets containing reduced inclusion of oil in the diet and without the use of growth-promoting antibiotics of broilers considering the effect on health, performance, and meat. Three treatments were defined: positive control (TP: with enramycin), negative control (TN: no enramycin), and blend (T-FRA: with monolaurin and glycerides of acid butyric minus 0.8% soybean oil). At 21 days, broilers treated with TP and T-FRA obtained the lower feed conversion ratio (FC); at 35 days, T-FRA broilers obtained lower FC than TN broilers. Cholesterol levels were higher in the blood of T-FRA broilers. On day 42, levels of ROS and TBARS were lower in the intestine, muscles, and liver of T-FRA broilers. Moreover, glutathione S-transferase and total non-enzymatic antioxidants were greater at the intestinal and muscular levels. The T-FRA broilers had a lower percentage of lipids in the meat. The MIC indicated that 111mg of the blend/mL inhibited the growth of E. coli; however, the counts of total coliforms and E. coli in the feces and the broilers' litter did not differ between treatments. In conclusion, the addition of the blend T-FRA in broiler diets was able to improve the feed conversion and maintain the other performance parameters even considering a reduction of 0.8% in the inclusion of oil.

New Pyridone-Based Derivatives as Cannabinoid Receptor Type 2 Agonists.

The activation of the human cannabinoid receptor type II (CB2R) is known to mediate analgesic and anti-inflammatory processes without the central adverse effects related to cannabinoid receptor type I (CB1R). In this work we describe the synthesis and evaluation of a novel series of N-aryl-2-pyridone-3-carboxamide derivatives tested as human cannabinoid receptor type II (CB2R) agonists. Different cycloalkanes linked to the N-aryl pyridone by an amide group displayed CB2R agonist activity as determined by intracellular [cAMP] levels. The most promising compound 8d exhibited a non-toxic profile and similar potency (EC50 = 112 nM) to endogenous agonists Anandamide (AEA) and 2-Arachidonoylglycerol (2-AG) providing new information for the development of small molecules activating CB2R. Molecular docking studies showed a binding pose consistent with two structurally different agonists WIN-55212-2 and AM12033 and suggested structural requirements on the pyridone substituents that can satisfy the orthosteric pocket and induce an agonist response. Our results provide additional evidence to support the 2-pyridone ring as a suitable scaffold for the design of CB2R agonists and represent a starting point for further optimization and development of novel compounds for the treatment of pain and inflammation.

Impact of macromolecular crowding on the mesomorphic behavior of lipid self-assemblies.

Using LAURDAN fluorescence we observed that water dynamics measured at the interface of DOPC bilayers can be differentially regulated by the presence of crowded suspensions of different proteins (HSA, IgG, Gelatin) and PEG, under conditions where the polymers are not in direct molecular contact with the lipid interface. Specifically, we found that the decrease in water dipolar relaxation at the membrane interface correlates with an increased fraction of randomly oriented (or random coil) configurations in the polymers, as Gelatin > PEG > IgG > HSA. By using the same experimental strategy, we also demonstrated that structural transitions from globular to extended conformations in proteins can induce transitions between lamellar and non-lamellar phases in mixtures of DOPC and monoolein. Independent experiments using Raman spectroscopy showed that aqueous suspensions of polymers exhibiting high proportions of randomly oriented conformations display increased fractions of tetracoordinated water, a configuration that is dominant in ice. This indicates a greater capacity of this type of structure for polarizing water and consequently reducing its chemical activity. This effect is in line with one of the tenets of the Association Induction Hypothesis, which predicts a long-range dynamic structuring of water molecules via their interactions with proteins (or other polymers) showing extended conformations. Overall, our results suggest a crucial role of water in promoting couplings between structural changes in macromolecules and supramolecular arrangements of lipids. This mechanism may be of relevance to cell structure/function when the crowded nature of the intracellular milieu is considered.

Synthesis of EPA- and DHA-Enriched Structured Acylglycerols at the sn-2 Position Starting from Commercial Salmon Oil by Enzymatic Lipase Catalysis under Supercritical Conditions.

There is consistent evidence that long-chain polyunsaturated fatty acids (LCPUFA) belonging to the n-3 series, i.e., eicosapentaenoic (20:5n-3, EPA) and docosahexaenoic (22:6n-3, DHA) acids, decrease the risk of heart, circulatory and inflammatory diseases. Furthermore, the bioavailability of such fatty acids has been shown to depend on their location in triacylglycerol (TG) molecules at the sn-2 position. Consequently, great attention has been accorded to the synthesis of structured acylglycerols (sAG), which include EPA or DHA at the sn-2 position. The aim of this work was to synthesize sAG starting from deodorized refined commercial salmon oil. For this, immobilized lipase B from Candida antarctica (nonspecific) was used as a catalyst for the intra-interesterification process under CO2 supercritical conditions (CO2SC). According to the CO2SC reaction time, three different fractions including sAG compounds were obtained. The location of EPA and DHA at the sn-2 position in the resulting glycerol backbone was identified by mass spectrometry (MALDI-TOF) analysis. In all fractions obtained, a marked decrease in the starting TG content was observed, while an increase in the DHA content at the sn-2 position was detected. The fraction obtained after the longest reaction time period (2 h) led to the highest yield of sn-2 position DHA in the resulting sAG molecule.

Protective role of endocannabinoid signaling in an animal model of haloperidol-induced tardive dyskinesia.

Tardive dyskinesia (TD) is a side effect associated with the long-term use of certain antipsychotics. Considering the modulatory role of the endocannabinoid system upon dopaminergic neurotransmission, the present study tested the hypothesis that increasing endocannabinoid (anandamide and 2-arachidonoylglycerol) levels attenuates haloperidol-induced TD (vacuous chewing movements, VCMs) in male Wistar rats. The animals received administration of chronic haloperidol (38 mg/kg; 29 days) followed by acute FAAH (URB597, 0.1-0.5 mg/kg) or MAGL (JZL184, 1-10 mg/kg) inhibitors before VCM quantification. The underlying mechanisms were evaluated by pre-treatments with a CB1 receptor antagonist (AM251, 1 mg/kg) or a TRPV1 channel blocker (SB366791, 1 mg/kg). Moreover, CB1 receptor expression was evaluated in the striatum of high-VCM animals. As expected, haloperidol induced VCMs only in a subset of rats. Either FAAH or MAGL inhibition reduced VCMs. These effects were prevented by CB1 receptor antagonism, but not by TRPV1 blockage. Remarkably, CB1 receptor expression was increased high-VCM rats, with a positive correlation between the levels of CB1 expression and the number of VCMs. In conclusion, increasing endocannabinoid levels results in CB1 receptor-mediated protection against haloperidol-induced TD in rats. The increased CB1 receptor expression after chronic haloperidol treatment suggests a counter-regulatory protective mechanism.

Cocaine-induced increases in motivation require 2-arachidonoylglycerol mobilization and CB1 receptor activation in the ventral tegmental area.

A wide body of evidence supports an integral role for mesolimbic dopamine (DA) in motivated behavior. In brief, drugs that increase DA in mesolimbic terminal regions, like cocaine, enhance motivation, while drugs that decrease DA concentration reduce motivation. Data from our laboratory and others shows that phasic activation of mesolimbic DA requires signaling at cannabinoid type-1 (CB1) receptors in the ventral tegmental area (VTA), and systemic delivery of CB1 receptor antagonists reduces DA cell activity and attenuates motivated behaviors. Recent findings demonstrate that cocaine mobilizes the endocannabinoid 2-arachidonoylglycerol (2-AG) in the VTA to cause phasic activation of DA neurons and terminal DA release. It remains unclear, however, if cocaine-induced midbrain 2-AG signaling contributes to the motivation-enhancing effects of cocaine. To examine this, we trained male and female rats on a progressive ratio (PR) task for a food reinforcer. Each rat underwent a series of tests in which they were pretreated with cocaine alone or in combination with systemic or intra-VTA administration of the CB1 receptor antagonist rimonabant or the 2-AG synthesis inhibitor tetrahydrolipstatin (THL). Cocaine increased motivation, measured by augmented PR breakpoints, while rimonabant dose-dependently decreased motivation. Importantly, intra-VTA administration of rimonabant or THL, at doses that did not decrease breakpoints on their own, blocked systemic cocaine administration from increasing breakpoints in male and female rats. These data suggest that cocaine-induced increases in motivation require 2-AG signaling at CB1 receptors in the VTA and may provide critical insight into cannabinoid-based pharmacotherapeutic targets for the successful treatment of substance abuse.

Targeting 2-arachidonoylglycerol signalling in the neurobiology and treatment of depression.

The endocannabinoid 2-arachidonoylglycerol (2-AG) is an atypical neurotransmitter synthesized on demand in response to a wide range of stimuli, including exposure to stress. Through the activation of cannabinoid receptors, 2-AG can interfere with excitatory and inhibitory neurotransmission in different brain regions and modulate behavioural, endocrine and emotional components of the stress response. Exposure to chronic or intense unpredictable stress predisposes to maladaptive behaviour and is one of the main risk factors involved in developing mood disorders, such as major depressive disorder (MDD). In this review, we describe the molecular mechanisms involved in 2-AG signalling in the brain of healthy and stressed animals and discuss how such mechanisms could modulate stress adaptation and susceptibility to depression. Furthermore, we review preclinical evidence indicating that the pharmacological modulation of 2-AG signalling stands as a potential new therapeutic target in treating MDD. Particular emphasis is given to the pharmacological augmentation of 2-AG levels by monoacylglycerol lipase (MAGL) inhibitors and the modulation of CB2 receptors.

Glyceryl monostearate-based oleogels as a new fat substitute in meat emulsion.

Bologna sausages were produced with 25, 50, 75 and 100% of their pork fat content replaced by monoglyceride based-oleogels prepared from conventional or high oleic sunflower oils. Physicochemical, technological, and sensory properties of Bologna sausages were evaluated. Emulsion stability was little affected by fat replacement. All treatments batters exhibited characteristic rheological properties of gels (G' > G″). Overall, the addition of oleogel as a fat substitute made the sausages lighter and a small increase in hardness was observed in the sausages with total fat replacement by oleogels. The sliceability was affected by the reformulation and a higher number of slices were obtained in samples with oleogels in relation to the control. These results were associated to the product structure that became more compact as the amount of pork fat was reduced. However, all samples showed good acceptance by the consumers and no significant difference was observed between treatments. The results showed that monostearate-based oleogel can be a potential fat replacer with higher amount of unsaturated fatty acids to be used in meat products, but retaining the desired characteristics of the traditional products.

Monoolein-based nanoparticles containing indinavir: a taste-masked drug delivery system.

OBJECTIVE: This study developed a novel child-friendly drug delivery system for pediatric HIV treatment: a liquid, taste-masked, and solvent-free monoolein-based nanoparticles formulation containing indinavir (0.1%). SIGNIFICANCE: Adherence to antiretroviral therapy by pediatric patients is difficult because of the lack of dosage forms adequate for children. METHODS: Monoolein-based nanoparticles were developed. The particle size, zeta potential, pH, drug content, small angle X-ray scattering, stability, in vitro drug release profile, biocompatibility, toxicity, and taste-masking properties were evaluated. RESULTS: Monoolein-based formulations containing indinavir had nanosized particles with 155 ± 7 nm, unimodal particle size distribution, and polydispersity index of 0.16 ± 0.03. The zeta potential was negative (-31.3 ± 0.3 mV) and pH was neutral (7.78 ± 0.01). A 96% drug incorporation efficiency was achieved, and the indinavir concentration remained constant for 30 days. Polarized light microscopy revealed isotropic characteristics. Transmission electron microscopy images showed spherical shaped morphology. Small-angle X-ray scattering displayed a form factor broad peak. Indinavir had a sustained release from the nanoparticles. The system was nonirritant and was able to mask drug bitter taste. CONCLUSIONS: Monoolein-based nanoparticles represent a suitable therapeutic strategy for antiretroviral treatment with the potential to reduce the frequency of drug administration and promote pediatric adherence.

Neem secretory cells: developmental cytology and indications of cell autotoxicity.

The neem tree (Azadirachta indica A.Juss.) contains a range of biologically active compounds-mainly triterpenoids produced in single secretory cells, which are distributed among all plant parts. Neem secretions are toxic to animal cells, triggering autolytic mechanisms that culminate in cell disruption. However, little is known about the self-toxicity of these secretions to the cells that produce them. We carried out an anatomical, histochemical, and ultrastructural investigation of neem's single secretory cells in the shoot apex and in young leaves. We evaluated the morphological changes as possible evidences of stress reactions to their own secretions. The subcellular apparatus involved in synthesis and compartmentation was consistent with hydrophilic and lipophilic secretions. Polymorphic plastids devoid of thylakoids and abundant smooth endoplasmic reticulum in the later stages of differentiation are comparable with previous reports on neem cotyledons with regard to terpenoid synthesis. However, secretions were compartmentalized within autophagic vacuoles and periplasmic spaces instead of in terpenoid vesicles. Cellular swelling, increased vesiculation, dilatation of endoplasmic reticulum cisternae, mitochondrial hypertrophy in the cristolysis process, autolytic vacuoles, and vacuolar degeneration culminating in protoplast autolysis are all consistent with early indications of autotoxicity. The signaling stress reaction mechanism was expressed as cytoplasmic deposits of calcium salt and by the expression of a 70-kDa heat-shock protein. The morphological and histochemical changes in the secreting cells are comparable with those described in animal cells exposed to neem oil. Our data provide evidence of cell damage and signaling reactions linked to these cells' own secretions before autolysis.

Microemulsion systems to enhance the transdermal permeation of ivermectin in dogs: A preliminary in vitro study.

This study aims to evaluate the influence of the phase behavior of microemulsions in the transdermal administration ("spot-on") of ivermectin, an antiparasitic drug widely used in the treatment of endoparasites and ectoparasites in dogs. In this regard, pseudoternary phase diagrams composed of water (aqueous phase), isopropyl myristate (oil phase), tween 80 (surfactant) and labrasol (cosurfactant) were obtained in a different surfactant: cosurfactant (S:CS) ratios. S:CS in 1:3 ratio presented a larger region of microemulsion formation and three microemulsions were selected from it and characterized. Subsequently, in vitro permeation and retention studies were conducted using canine skin as membrane. SAXS, rheology and conductivity data were employed to confirm the phase behavior of the microemulsions (w/o, bicontinuous or o/w). The cutaneous permeation and retention tests showed that the w/o microemulsion, followed by bicontinuous microemulsion, resulted in a higher amount of drug permeated through canine skin, suggesting better transdermal permeation. On the other hand, o/w microemulsion resulted in a higher amount of drug accumulated into the skin, suggesting better topical activity. Thus, it can be concluded that phase behavior of microemulsions influenced the drug permeation in the canine skin differently from other animal models. Microemulsions, especially w/o and bicontinuous, can be promising vehicles regarding the transdermal delivery of ivermectin.

A micro salting-out assisted liquid-liquid extraction combined with ultra-high performance liquid chromatography tandem mass spectrometry to determine anandamide and 2-arachidonoylglycerol in rat brain samples.

A simple and reliable method was developed and validated to determine the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG) in rat brain samples by micro salting-out assisted liquid-liquid extraction combined with ultra-high performance liquid chromatography tandem mass spectrometry (SALLLE/UHPLC-MS/MS). The SALLE parameters (brain homogenate volume, salting-out agent, salt concentration, salt solution volume, organic solvent, organic solvent volume, and centrifugation temperature) were optimized to improve sensitivity and selectivity of the method. The SALLE/UHPLC-MS/MS method presented linear ranges from 2.00 to 20.00 ng mL-1 for AEA and from 0.300 to 10.00 µg mL-1 for 2-AG, no significant matrix effect, and inter- and intra-assay precision and accuracy with CV and RSE values lower than 15%, respectively. This innovative method was successfully applied to determine AEA and 2-AG in brain hemispheres from a 6-OHDA animal model of Parkinson's disease (PD).