Scrubbed Palm Fatty Acid Distillate as Vitamin E Concentrate.

Vitamin E (VitE) production from crude palm oil (CPO) has been extensively studied and industrially conducted. VitE in CPO is in the range of 600 to 1,000 ppm, and is usually produced from one of the main by-products of edible palm oil production, namely palm fatty acid distillate (PFAD). PFAD contains 4,000 to 5,500 ppm of VitE, and is produced from deodorization process of palm oil purification. This paper presents an innovative process of VitE concentrate production from CPO. A scrubber was designed and installed between the deodorizer and conventional PFAD scrubber. The main objective of this new scrubber was a recovery of glycerides from PFAD. This new scrubber is operated at 150 to 160℃. The scrubbed oil is named as Scrubbed Palm Fatty Acid Distillate (S-PFAD). This simple and efficient modified process can retrieve glycerides as S-PFAD at 0.3% recovery and it enhances VitE concentration in S-PFAD to the range of 28,000 to 32,000 ppm, which is the highest concentration of VitE that has ever been produced in the palm oil production. Fatty acids and glycerides in S-PFAD were esterified and transesterified to methyl esters. The methyl esters were evaporated from S-PFAD, and S-PFAD residue oil contained 24.7% VitE.

In-tube solid-phase microextraction directly coupled to tandem mass spectrometry for anandamide and 2-arachidonoylglycerol determination in rat brain samples from an animal model of Parkinson's disease.

To evaluate the endocannabinoid system in an animal model of Parkinson's disease, in-tube solid-phase microextraction (in-tube SPME) was directly coupled to a tandem mass spectrometry (MS/MS) system for determination of the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG) in rat brain samples. In-tube SPME-which consisted of a microtube of restricted access material (RAM) with a hydrophilic diol external surface and a hydrophobic octyl inner surface-efficiently excluded (up to 95%) macromolecules from the biological samples and selectively pre-concentrated the analytes. In-tube SPME parameters, such as sample volume, mobile phases, flow rate, and pre-concentration time, were evaluated to improve the extraction efficiency and throughput performance. The selectivity of the in-tube SPME and MS/MS (MRM mode) techniques allowed them to be directly coupled online, which dismissed the need for the chromatographic separation step. The in-tube SPME-MS/MS method was validated and shown to be linear from 6.0 to 30.0 ng mL-1 for AEA and from 10.0 to 100.0 ng mL-1 for 2-AG; the intra- and inter-assay accuracy and precision were lower than 15%. Parallelism between the calibration curves constructed in the matrix and aqueous solution confirmed that there was no matrix effect. The method allowed endogenous concentrations of AEA and 2-AG to be determined in rat brain striatum from unilaterally 6-hydroxydopamine-lesioned animals. The concentrations of these endocannabinoids in striatum ipsilateral and contralateral to the lesion differed significantly (p<0.001).

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).

A normal phase high performance liquid chromatography method for the separation of hydroxy and non-hydroxy neutral lipid classes compatible with ultraviolet and in-line liquid scintillation detection of radioisotopes.

In this paper, we report a method for the separation of hydroxy fatty acid and non-hydroxy fatty acid containing neutral lipid classes via normal phase HPLC with UV detection on a PVA-Sil column. The hexane/isopropanol/methanol/water based method separates all the neutral lipids in 21 min, and subsequently flushes through the polar lipids by 27 min such that prefractionation of neutral and polar lipids are not required, and the column is re-equilibrated for the next run in 15 min, for a total run time of 45 min per sample. The separation was demonstrated at both 1.0 mL/min and 1.5 mL/min for added applicability for fraction collection or inline analysis. Separation of various hydroxy fatty acid containing lipids was demonstrated from three different plant species Ricinus communis, Physaria fendleri, and engineered Arabidopsis thaliana. Additionally, we have combined this method with an in-line liquid scintillation counter for the separation and quantification of 14C labeled lipids obtained from in vivo metabolic flux experiments conducted in the developing seeds of Arabidopsis thaliana.

Isolation and Identification of Benzochroman and Acylglycerols from Massa Medicata Fermentata and Their Inhibitory Effects on LPS-Stimulated Cytokine Production in Bone Marrow-Derived Dendritic Cells.

Massa Medicata Fermentata (MMF), known as Shenqu, is an important traditional Chinese medicine widely used to treat indigestion, vomiting, and diarrhea. In this study, a new benzochroman, 3(S)-3,4-dihydro-5,10-di-ß-d-glucopyranoside-2,2-dimethyl-2H-naphtho(2,3-b)pyran-3-ol (1), and five known galactosyl acylglycerols (2⁻6) were isolated from a methanol extract from MMF. In addition, their chemical structures were determined by chemical and spectroscopic methods, which were compared with the previously reported data. Furthermore, the effects of isolated compounds on lipopolysaccharide (LPS)-stimulated bone marrow-derived dendritic cells were investigated. Compounds 1⁻3 exhibited significant inhibitory effects on the LPS-induced production of IL-6 and IL-12 p40, with IC50 values ranging from 1.6 to 10.2 µM. Compounds 2 and 3 also exhibited strong inhibitory effects on the LPS-stimulated production of TNF-α with IC50 values of 12.0 and 11.2 µM, respectively. The results might provide a scientific basis for the development of the active components in MMF, as well as for novel anti-inflammatory agents.

Glycosyl glycerides from the aerial parts of Malva verticillata and their chemopreventive effects.

A new glycosyl glyceride (5) along with twelve known ones (1-4 and 6-13) including two sulfoquinovosyl glycerides (1 and 2) were isolated from the aerial parts of Malva verticillata. Based on several spectroscopic methods, compound 5 was identified to be (2S)-1-O-ß-d-galactopyranosyl-3-O-isostearoyl glyceride, and named malvaglycolipid A. Compounds 1 and 2 contained a unique sugar, (6-deoxy-6-sulfo)-α-d-glucopyranose, which very rarely occurs in natural sources. This is the first report for the isolation of compounds 1 and 2 from natural sources and the structure determination using NMR experiment. It was also of note that no glycosyl glyceride has previously been isolated from the family of Malvaeae. Most glycosyl glycerides showed cytotoxicity to HepG2, AGS, HCT-15, and A549 human cancer cells. Especially, compounds 1, 2, and 11 exhibited significant cytotoxicity to AGS cells, with IC50 values of 33.7 ±â€¯0.64 µM, 11.1 ±â€¯0.07 µM, and 10.6 ±â€¯0.10 µM, respectively. The n-BuOH fraction and compounds 1, 2, and 11 increased the number of apoptotic cells in the Tali assay and had a significant effect on the levels of proteins related to apoptosis including PARP, caspase-3, Bcl-2, Bax, and ß-actin.

Inflammation Modulatory Phorbol Esters from the Seeds of Aquilaria malaccensis.

The tree Aquilaria malaccensis is a valuable source of agarwood, which is used in herbal medicinal preparations. Phytochemical research on A. malaccensis seeds has led to the isolation of four new phorbol esters (1-4), two known phorbol esters (5, isolated from Nature for the first time, and 6), and two known glycerides (7 and 8). The structures of these isolates were elucidated by means of spectroscopic data interpretation. The inflammation-modulatory activities of the isolates on elastase release and superoxide anion generation in human neutrophils were evaluated. Interestingly, phorbol esters 1, 5, and 6 showed potent inhibitory activity on elastase release in human neutrophils, with IC50 values of 2.7, 0.8, and 2.1 µM, respectively. All isolated phorbol esters exerted enhancing activity on superoxide anion generation. The results indicated that phorbol esters may play a bilateral modulatory role in the processes of inflammation. In addition, the compounds were evaluated for their cytotoxic properties against HepG2 (hepatoma), MDA-MB-231 (breast), and A549 (lung) cancer cells, but all compounds were inactive for all cell lines used (IC50 > 10 µM).

Stabilization of Neem Oil Biodiesel with Corn Silk Extract during Long-term Storage.

The current study aimed to evaluate the antioxidant efficiency of different extracts of corn silk. In addition, the impact of corn silk extract on oxidative stability of neem biodiesel during storage was studied. The highest phenolics, DPPH radical scavenging and reducing power activities were recorded for methanol-water extract. The longest oxidation stability (10 h) was observed for biodiesel samples blended with 1000 ppm of corn silk extract (CSE). At the end of storage period the induction time of biodiesel samples mixed with 1000 ppm of CSE or butylated hydroxytoluene (BHT) were about 6.72 and 5.63 times as high as in biodiesel samples without antioxidants. Biodiesel samples blended with 1000 ppm of CSE had the lowest acidity at the end of storage period. Peroxide value of biodiesel samples containing 1000 ppm of CSE was about 4.28 times as low as in control sample without antioxidants.

Mirabolides A and B; New Cytotoxic Glycerides from the Red Sea Sponge Theonella mirabilis.

As a part of our continuing work to find out bioactive lead molecules from marine invertebrates, the CHCl3 fraction of the organic extract of the Red Sea sponge Theonella mirabilis showed cytotoxic activity in our primary screen. Bioassay-guided purification of the active fractions of the sponge's extract resulted in the isolation of two new glycerides, mirabolides A and B (1 and 2), together with the reported 4-methylene sterols, conicasterol (3) and swinhosterol B (4). The structures of the compounds were assigned by interpretation of their 1D (¹H, (13)C), 2D (COSY, HSQC, HMBC, ROESY) NMR spectral data and high-resolution mass determinations. Compounds 1-4 displayed marked cytotoxic activity against human breast adenocarcinoma cell line (MCF-7) with IC50 values of 16.4, 5.18, 6.23 and 3.0 µg/mL, respectively, compared to 5.4 µg/mL observed by doxorubicin as reference drug.

Determination of 2-Arachidonoylglycerol by µSPE-LC-MS/MS.

LC-MS/MS is a powerful analytical technique that provides unequivocal identification and reliable quantification of the analytes, using Selected Reaction Monitoring or Multi Reaction Monitoring acquisition mode.2-Arachidonoylglycerol (2-AG) is the most abundant endocannabinoid, which plays a major role in a wide variety of physiological and pathological processes. Analysis of 2-AG by means of LC-MS/MS allows the detection of very low concentrations in biological samples. Here, we describe how to determine 2-AG levels in tiny samples of tissues and plasma through LC-MS/MS, by using very quick and easy to perform extraction procedures, with reduced solvent consumption.

High-performance liquid chromatography - Ultraviolet method for the determination of total specific migration of nine ultraviolet absorbers in food simulants based on 1,1,3,3-Tetramethylguanidine and organic phase anion exchange solid phase extraction to remove glyceride.

The glyceride in oil food simulant usually causes serious interferences to target analytes and leads to failure of the normal function of the RP-HPLC column. In this work, a convenient HPLC-UV method for the determination of the total specific migration of nine ultraviolet (UV) absorbers in food simulants was developed based on 1,1,3,3-tetramethylguanidine (TMG) and organic phase anion exchange (OPAE) SPE to efficiently remove glyceride in olive oil simulant. In contrast to the normal ion exchange carried out in an aqueous solution or aqueous phase environment, the OPAE SPE was performed in the organic phase environments, and the time-consuming and challenging extraction of the nine UV absorbers from vegetable oil with aqueous solution could be readily omitted. The method was proved to have good linearity (r≥0.99992), precision (intra-day RSD≤3.3%), and accuracy(91.0%≤recoveries≤107%); furthermore, the lower limit of quantifications (0.05-0.2mg/kg) in five types of food simulants(10% ethanol, 3% acetic acid, 20% ethanol, 50% ethanol and olive oil) was observed. The method was found to be well suited for quantitative determination of the total specific migration of the nine UV absorbers both in aqueous and vegetable oil simulant according to Commission Regulation (EU) No. 10/2011. Migration levels of the nine UV absorbers were determined in 31 plastic samples, and UV-24, UV-531, HHBP and UV-326 were frequently detected, especially in olive oil simulant for UV-326 in PE samples. In addition, the OPAE SPE procedure was also been applied to efficiently enrich or purify seven antioxidants in olive oil simulant. Results indicate that this procedure will have more extensive applications in the enriching or purification of the extremely weak acidic compounds with phenol hydroxyl group that are relatively stable in TMG n-hexane solution and that can be barely extracted from vegetable oil.

Chemical constituents and antiproliferative effects of cultured Mougeotia nummuloides and Spirulina major against cancerous cell lines.

In this study, the effect of Mougeotia nummuloides and Spirulina major on Vero cells (African green monkey kidney), C6 cells (rat brain tumor cells) and HeLa cells (human uterus carcinoma) was investigated in vitro. The antiproliferative effect of the methanol extract of M. nummuloides and S. major compared with 5-fluorourasil (5-FU) and cisplatin was tested at various concentrations using the BrdU Cell Proliferation ELISA. Both M. nummuloides and S. major extracts significantly inhibited the proliferation of Vero, HeLa and C6 cancer cell lines with IC50 and IC75 values. The M. nummuloides extract exhibited higher activity than 5-FU and cisplatin on Vero and C6 cells at high concentrations. The S. major extract revealed better antifproliferative activity than standards against Vero cells at 500 µg/mL. The compounds of methanol extracts were determined by GC-MS after the silylation process. Trehalose, monostearin and 1-monopalmitin were detected as major products in the M. nummuloides extract where as in the S. major extract; monostearin, 1-monopalmitin and hexyl alcohol were the main constituents.

Anti-MRSA and anti-TB metabolites from marine-derived Verrucosispora sp. MS100047.

Microbes belonging to the genus Verrucosispora possess significant chemical diversity and biological properties. They have attracted the interests of many researchers and are becoming promising resources in the marine natural product research field. A bioassay-guided isolation from the crude extract of Verrucosispora sp. strain MS100047, isolated from sediments collected from the South China Sea, has led to the identification of a new salicylic derivative, glycerol 1-hydroxy-2,5-dimethyl benzoate (1), along with three known compounds, brevianamide F (2), abyssomicin B (3), and proximicin B (4). Compound 1 showed selective activity against methicillin-resistant Staphylococcus aureus (MRSA) with a minimum inhibitory concentration (MIC) value of 12.5 µg/mL. Brevianamide F (2), which was isolated from actinomycete for the first time, showed a good anti-BCG activity with a MIC value of 12.5 µg/mL that has not been reported previously in literatures. Proximicin B (4) showed significant anti-MRSA (MIC = 3.125 µg/mL), anti-BCG (MIC = 6.25 µg/mL), and anti-tuberculosis (TB) (MIC = 25 µg/mL) activities. This is the first report on the anti-tubercular activities of proximicins. In addition, Verrucosispora sp. strain MS100047 was found to harbor 18 putative secondary metabolite gene clusters based on genomic sequence analysis. These include the biosynthetic loci encoding polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) consistent with abyssomicins and proximicins, respectively. The biosynthetic pathways of these isolated compounds have been proposed. These results indicate that MS100047 possesses a great potential as a source of active secondary metabolites.

Selective production of 1-monocaprin by porcine liver carboxylesterase-catalyzed esterification: Its enzyme kinetics and catalytic performance.

Porcine liver carboxylesterase (PLE) belongs to carboxylesterase family (EC 3.1.1.1) as a serine-type esterase. The PLE-catalyzed esterification of capric acid with glycerol in reverse micelles was investigated on the catalytic performance and enzyme kinetics. The most suitable structure of reverse micelles was comprised of isooctane (reaction medium) and bis(2-ethylhexyl) sodium sulfosuccinate (AOT, anionic surfactant) with 0.1 of R-value ([water]/[surfactant]) and 3.0 of G/F-value ([glycerol]/[fatty acid]) for the PLE-catalyzed esterification. In the aspect of regio-selectivity, the PLE mainly produced 1-monocaprin without any other products (di- and/or tricaprins of subsequent reactions). Furthermore, the degree of esterification at equilibrium state (after 4 h from the initiation) was 62.7% under the optimum conditions at pH 7.0 and 60 °C. Based on Hanes-Woolf plot, the apparent Km and Vmax values were calculated to be 16.44 mM and 38.91 µM/min/mg protein, respectively.

Old ingredients for a new recipe? Neem cake, a low-cost botanical by-product in the fight against mosquito-borne diseases.

Mosquitoes (Diptera: Culicidae) represent an important threat to millions of people worldwide, since they act as vectors for important pathogens, such as malaria, yellow fever, dengue and West Nile. Control programmes mainly rely on chemical treatments against larvae, indoor residual spraying and insecticide-treated bed nets. In recent years, huge efforts have been carried out to propose new eco-friendly alternatives, with a special focus on the evaluation of plant-borne mosquitocidal compounds. Major examples are neem-based products (Azadirachta indica A. Juss, Meliaceae) that have been proven as really effective against a huge range of pests of medical and veterinary importance, including mosquitoes. Recent research highlighted that neem cake, a cheap by-product from neem oil extraction, is an important source of mosquitocidal metabolites. In this review, we examined (i) the latest achievements about neem cake metabolomics with special reference to nor-terpenoid and related content; (ii) the neem cake ovicidal, larvicidal and pupicidal toxicity against Aedes, Anopheles and Culex mosquito vectors; (iii) its non-target effects against vertebrates; and (iv) its oviposition deterrence effects on mosquito females. Overall, neem cake can be proposed as an eco-friendly and low-cost source of chemicals to build newer and safer control tools against mosquito vectors.

Truffles contain endocannabinoid metabolic enzymes and anandamide.

Truffles are the fruiting body of fungi, members of the Ascomycota phylum endowed with major gastronomic and commercial value. The development and maturation of their reproductive structure are dependent on melanin synthesis. Since anandamide, a prominent member of the endocannabinoid system (ECS), is responsible for melanin synthesis in normal human epidermal melanocytes, we thought that ECS might be present also in truffles. Here, we show the expression, at the transcriptional and translational levels, of most ECS components in the black truffle Tuber melanosporum Vittad. at maturation stage VI. Indeed, by means of molecular biology and immunochemical techniques, we found that truffles contain the major metabolic enzymes of the ECS, while they do not express the most relevant endocannabinoid-binding receptors. In addition, we measured anandamide content in truffles, at different maturation stages (from III to VI), through liquid chromatography-mass spectrometric analysis, whereas the other relevant endocannabinoid 2-arachidonoylglycerol was below the detection limit. Overall, our unprecedented results suggest that anandamide and ECS metabolic enzymes have evolved earlier than endocannabinoid-binding receptors, and that anandamide might be an ancient attractant to truffle eaters, that are well-equipped with endocannabinoid-binding receptors.

Statistical Optimization of the Content Composition Precursors Using Response Surface Methodology to Enhance Agaricoglyceride A Production from the Shaggy Ink Cap Medicinal Mushroom, Coprinus comatus (Higher Basidiomycetes) Mycelia.

Coprinus comatus, a novel cultivated edible mushroom, has a various of pharmacological effects due to its many active components. In this study, agaricoglycerides, a new class of fungal secondary metabolites that have strong activity against neurolysin, were isolated from C. comatus mycelia. Simultaneously, a 3-level Box-Behnken factorial design was used, combined with response surface methodology, to optimize the precursor composition of agaricoglycerides for the production of agaricoglyceride A. The model estimated that a maximal yield of agaricoglyceride A (20.105 mg/L) could be obtained when the concentrations of 4-hydroxybenzoic acid, glycerol, and methanol (MeOH) were set at 75 mg/L, 0.75 mL/L, and 0.75 mL/L, respectively. The verified experiments showed that the model was significantly consistent with the model prediction. These results showed that appropriately adding the precursors could increase the production of agaricoglyceride A.

Didemnacerides A and B: two new glycerides from Red Sea ascidian Didemnum species.

Two new glycerides, didemnacerides A (1) and B (2), together with three known sterols, 24-ethyl-25-hydroxycholesterol (3), cholest-6-en-3,5,8-triol (4) and cholestane-3ß,5α,6ß-26-tetrol (5), were isolated from the Red Sea ascidian Didemnum sp. Their structures were elucidated by using extensive 1D ((1)H, (13)C) and 2D ((1)H-(1)H COSY, HSQC and HMBC) NMR studies and mass spectroscopic data (GC-MS and HR-MS) as well as alkaline hydrolysis followed by GC-MS and NMR spectral analyses of the fatty acid methyl esters. This is the first report of compounds 3-5 from the Red Sea ascidian Didemnum species.

Anti-inflammatory potential of monogalactosyl diacylglycerols and a monoacylglycerol from the edible brown seaweed Fucus spiralis Linnaeus.

A monoacylglycerol (1) and a 1:1 mixture of two monogalactosyl diacylglycerols (MGDGs) (2 and 3) were isolated from the brown seaweed Fucus spiralis Linnaeus. The structures were elucidated by spectroscopic means (NMR and MS) and by comparison with the literature. Compound 1 was composed of a glycerol moiety linked to oleic acid (C18:1 Ω9). Compounds 2 and 3 contained a glycerol moiety linked to a galactose unit and eicosapentaenoic acid (C20:5 Ω3) combined with octadecatetraenoic acid (C18:4 Ω3) or linolenic acid (C18:3 Ω3), respectively. The isolated compounds were tested for their cytotoxic and anti-inflammatory activity in RAW 264.7 macrophage cells. All of them inhibited NO production at non-cytotoxic concentrations. The fraction consisting of compounds 2 and 3, in a ratio of 1:1, was slightly more effective than compound 1 (IC50 of 60.06 and 65.70 µg/mL, respectively). To our knowledge, this is the first report of these compounds from F. spiralis and on their anti-inflammatory capacity.

Adsorption of glycerol, monoglycerides and diglycerides present in biodiesel produced from soybean oil.

The most common methods currently used for the removal of waste glycerol, monoglycerides and diglycerides remaining after phase separation during biodiesel production involve wet processes. These procedures are not environmentally viable because they require large volumes of water and thus generate significant quantities of effluent. In this study, adsorption was employed to replace this purification step. Some commercial activated carbons were tested along with adsorbents chemically modified with HNO3. A kinetics study was conducted at 30 degrees C and adsorption isotherms were obtained at 20 degrees C, 30 degrees C and 40 degrees C. The results indicated that the adsorption of glycerol increased with the use of chemically-modified activated carbon, showing that pH has a strong influence on glycerol adsorption. The pseudo-first-order kinetic model provided the best fit with the experimental data for the monoglycerides while the pseudo-second-order model showed a better fit for the glycerol and diglycerides. The Freundlich model had the best fit with experimental data on the adsorption equilibrium for all temperatures. The thermodynamic study indicated that the adsorption process is endothermic and thus adsorption is favoured by increasing the temperature. The adsorption process using chemically-modified activated carbon was therefore very effective for the removal of waste glycerol resulting from biodiesel production, which is of considerable significance given the legal limits imposed.