Foxq1 activates CB2R with oleamide to alleviate POCD.

Postoperative cognitive dysfunction (POCD) is a major concern, particularly among older adults. This study used social isolation (ISO) and multiomics analyses in aged mice to investigate potential mechanisms underlying POCD development. Aged mice were divided into two groups: ISO and paired housing (PH). Oleamide and the cannabinoid receptor type 2 (CB2R) antagonist AM630 were administered intraperitoneally, while Foxq1 adeno-associated viral (AAV) vector was injected directly into the hippocampus. Intramedullary tibial surgeries were subsequently performed to establish the POCD models. Behavioral tests comprising the Y-maze, open field test, and novel object recognition were conducted 2 days after surgery. Hippocampal and serum inflammatory cytokines were assessed. Following surgery, ISO mice demonstrated intensified cognitive impairments and escalated inflammatory markers. Integrative transcriptomic and metabolomic analysis revealed elevated oleamide concentrations in the hippocampus and serum of PH mice, with associative investigations indicating a close relationship between the Foxq1 gene and oleamide levels. While oleamide administration and Foxq1 gene overexpression substantially ameliorated postoperative cognitive performance and systemic inflammation in mice, CB2R antagonist AM630 impeded these enhancements. The Foxq1 gene and oleamide may be crucial in alleviating POCD. While potentially acting through CB2R-mediated pathways, these factors may modulate neuroinflammation and attenuate proinflammatory cytokine levels within the hippocampus, substantially improving cognitive performance postsurgery. This study lays the groundwork for future research into therapeutic approaches targeting the Foxq1-oleamide-CB2R axis, with the ultimate goal of preventing or mitigating POCD.

Milk-based culture of Penicillium camemberti and its component oleamide affect cognitive function in healthy elderly Japanese individuals: a multi-arm randomized, double-blind, placebo-controlled study.

Dairy products and fermented foods have a reported association with maintained cognitive function. Camembert cheese, a dairy product fermented by the white mold Penicillium camemberti, has also been shown to enhance cognitive function in vivo. Oleamide, derived from the fermentation of the white mold, is a candidate for an active component, and expected to improve both cognitive function and sleep conditions. Thus, this study investigated whether the milk-based culture of white mold (MCW), and oleamide, could improve cognitive function and sleep state clinically. A multi-arm randomized, double-blind, placebo-controlled trial was conducted in Tokyo, Japan. 60 healthy Japanese individuals aged 50-75 who were aware of their cognitive decline were randomly and equally divided into three groups of 20 participants using computer-generated random numbers. Participants took either MCW (equivalent to 60 µg/day of oleamide), 60 µg/day of oleamide, or placebo capsules for 12 weeks. Serum BDNF, cognitive function by Cognitrax as primary and MCI Screen as secondary outcome, and sleep status using the Japanese version of the Pittsburgh Sleep Quality Index (PSQI-J) were assessed before and after intervention. The participants, outcome assessors and analysts, and research assistants were blinded to the group assignment. Of the 60 participants, 58 completed the study and were analyzed. No adverse events related to test foods were observed. The placebo group showed a negative rate of change in serum BDNF (-10.5% ± 19.7%), whereas the MCW and oleamide groups showed positive changes (2.0% ± 27.1% and 1.3% ± 13.5%, respectively). Cognitrax scores increased after 12 weeks in all groups. Conversely, the MPI score of the MCI Screen demonstrated a significant improvement in the MCW and oleamide groups compared to the placebo group (p = 0.013 and p < 0.001, respectively). The subscales, immediate free recall and delayed free recall, also significantly increased in them compared to the placebo group. Although PSQI-J revealed no significant differences among groups, the MCW and oleamide groups showed significant improvement after intervention in overall score, subjective sleep quality, and sleep latency. Our results suggest that MCW and its component, oleamide, are safe and contribute to maintaining cognitive functions, particularly short-term and working memory, and improving sleep state. Clinical trial registration: https://center6.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000054792, identifier UMIN-CTR UMIN000048084.

Pharmacometabolomic study of drug response to antihypertensive medications for hypertension marker identification in Han Chinese individuals in Taiwan.

Various groups of antihypertensive drugs targeting different pathways have been developed; however, the pharmacometabolic responses to these drugs have rarely been compared to elucidate the common pathway of blood pressure regulation. Here, we performed a comparative multi-dimensional pharmacometabolic study on the four major lines of antihypertensive drugs, namely angiotensin-converting enzyme inhibitors (ACEis), angiotensin receptor blockers (ARBs), calcium channel blockers (CCBs), and diuretics (DIURs), through ultra-performance liquid chromatography coupled to quantum time-of-flight mass spectrometry. Two hundred fifty patients with young-onset hypertension, who were equally divided among five study groups: non-medicated, ACEi, ARB, CCB, and DIUR groups, were recruited. In a metabolome-wide association study conducted through analysis of covariance, 37 molecular features significantly associated with pharmacometabolic responses to antihypertensive drugs were identified. One-third of these features were shared by multiple medications. ACEis, ARBs, and DIURs shared more features than CCB, partially reflecting that ACEis, ARBs, and DIURs affect the renin-angiotensin-aldosterone system. Thirteen molecular features were consistently identified by all four models of the analysis of covariance. A tandem mass spectrometry (or MS/MS) experiment was performed to decipher the chemical structure of these 13 molecular features, including ARB-associated lysophosphatidylcholine (P4135), CCB-associated diacylglycerol(15:0/18:2) (P1175), and DIUR-associated oleamide (P1516). In addition, diacylglycerol(15:0/14:2) (P408) was significantly associated with the pharmacometabolic response to all four antihypertensive drugs. The identified metabolites provide insights into the mechanisms of blood pressure regulation and potential predictive markers of pharmacometabolic responses to antihypertensive drugs.

Transport Form and Pathway from the Intestine to the Peripheral Tissues and the Intestinal Absorption and Metabolism Properties of Oleamide.

This study aimed to obtain information on the transport form and pathway from the intestine to the peripheral tissues and on the intestinal absorption and metabolism properties of oleamide (cis-9-octadecenamide). Oleamide was primarily transported via the portal vein. Density gradient centrifugation indicated that plasma oleamide was enriched in the fractions containing albumin in the portal and peripheral blood. Oleamide formed a complex with albumin in an endothermic reaction (apparent Kd = 4.4 µM). The CD36 inhibitor inhibited the oleamide uptake into the intestinal epithelial Caco-2 cells, and oleamide decreased the cell surface CD36 level. The fatty acid amide hydrolase (FAAH) inhibitor increased the transepithelial transport of oleamide across Caco-2 cells and the plasma oleamide concentration in mice intragastrically administered with oleamide. These results indicate that oleamide is transported primarily via the portal vein as a complex with albumin. Furthermore, we suggest that oleamide is taken up via CD36 in the small intestine and degraded intracellularly by FAAH.

Phytochemical and Antioxidant Profile of the Medicinal Plant Melia azedarach Subjected to Water Deficit Conditions.

Environmental stress triggered by climate change can alter the plant's metabolite profile, which affects its physiology and performance. This is particularly important in medicinal species because their economic value depends on the richness of their phytocompounds. We aimed to characterize how water deficit modulated the medicinal species Melia azedarach's lipophilic profile and antioxidant status. Young plants were exposed to water deficit for 20 days, and lipophilic metabolite profile and the antioxidant capacity were evaluated. Leaves of M. azedarach are rich in important fatty acids and oleamide. Water deficit increased the radical scavenging capacity, total phenol, flavonoids, and catechol pools, and the accumulation of ß-sitosterol, myo-inositol, succinic acid, sucrose, d-glucose and derivatives, d-psicofuranose, d-(+)-fructofuranose, and the fatty acids stearic, α-linolenic, linoleic and palmitic acids. These responses are relevant to protecting the plant against climate change-related stress and also increase the nutritional and antioxidant quality of M. azedarach leaves.

Oleamide Reduces Mitochondrial Dysfunction and Toxicity in Rat Cortical Slices Through the Combined Action of Cannabinoid Receptors Activation and Induction of Antioxidant Activity.

The potential treatment of neurodegenerative disorders requires the development of novel pharmacological strategies at the experimental level, such as the endocannabinoid-based therapies. The effects of oleamide (OEA), a fatty acid primary amide with activity on cannabinoid receptors, was tested against mitochondrial toxicity induced by the electron transport chain complex II inhibitor, 3-nitropropionic acid (3-NP), in rat cortical slices. OEA prevented the 3-NP-induced loss of mitochondrial function/cell viability at a concentration range of 5 nM-25 µM, and this protective effect was observed only when the amide was administered as pretreatment, but not as post-treatment. The preservation of mitochondrial function/cell viability induced by OEA in the toxic model induced by 3-NP was lost when the slices were pre-incubated with the cannabinoid receptor 1 (CB1R) selective inhibitor, AM281, or the cannabinoid receptor 2 (CB2R) selective inhibitor, JTE-907. The 3-NP-induced inhibition of succinate dehydrogenase (mitochondrial Complex II) activity was recovered by 25 nM OEA. The amide also prevented the increased lipid peroxidation and the changes in reduced/oxidized glutathione (GSH/GSSG) ratio induced by 3-NP. The cell damage induced by 3-NP, assessed as incorporation of cellular propidium iodide, was mitigated by OEA. Our novel findings suggest that the neuroprotective properties displayed by OEA during the early stages of damage to cortical cells involve the converging activation of CB1R and CB2R and the increase in antioxidant activity, which combined may emerge from the preservation of the functional integrity of mitochondria.

Oleamide-Mediated Polarization of M1 Macrophages and IL-1ß Production by Regulating NLRP3-Inflammasome Activation in Primary Human Monocyte-Derived Macrophages.

Macrophages are a type of innate immune cell that activates the NLRP3 inflammasome, causing the release of the cytokine IL-1ß, which is a crucial mediator of the inflammatory response. NLRP3 activation that is dysregulated worsens a variety of inflammatory and autoimmune diseases, as well as neurodegenerative diseases. Oleamide is an endogenous fatty acid amide that was first determined as a sleep-inducing molecule and later shown to have wide-ranging beneficial effects on the central nervous system. How oleamide influences human macrophage polarization and NLRP3-inflammasome activation remains unclear. The effect of oleamide on macrophage polarization was explored using an in vitro culture of primary human monocyte-derived macrophages (MDMs) supplemented with human serum-containing media. Cellular and molecular mechanisms of oleamide-regulated MDMs polarization were also investigated. Results showed that oleamide promoted naïve macrophages (M0) toward the M1 phenotype by upregulating M1-associated genes (IL-1ß, iNOS, CXCL10), along with downregulation of M2-associated genes (Arg-1, CD206, CCL22). Cell surface expression indicated that oleamide enhanced CD80 expression in M0 naïve macrophages and hider CD206 and CD163 expression in M2 macrophages. Higher production of IL-1ß cytokine was observed but with no alteration in IL-6 and TNF-α levels by MDMs and differentiated THP-1 models. Whether oleamide functioned as a second signal that activated the NLRP3 inflammasome and mediated IL-1ß production was further investigated using LPS-primed MDMs followed by oleamide treatment that induced activation of inflammasome-related proteins including NLRP3, ASC, cleaved casp-1, and cleaved IL-1ß. These findings suggested that oleamide promoted M1 macrophage polarization and increased IL-1ß production by activating the NLRP3 inflammasome in primary MDMs. This research reveals a new function for oleamide as well as prospective targets for treating NLRP3-related inflammatory disorders.

Dietary oleamide attenuates obesity induced by housing mice in small cages.

Physical inactivity due to prolonged sedentary behavior induces obesity. Therefore, we investigated whether housing mice in small cages to mimic sedentary behavior induced obesity and whether dietary oleamide (cis-9,10-octadeceneamide) suppressed the induced obesity. A single oral administration of oleamide (50 mg/kg) to mice resulted in the accumulation of the exogenous oleamide in abdominal visceral fat. Next, mice were housed in small cages and oleamide (50 mg/kg/d) was orally administered for 12 weeks. Housing mice in small cages impaired glucose tolerance and increased food efficiency. It also increased body weight and abdominal fat mass. Dietary oleamide improved the impairment and inhibited their increase in mice housed in small cages. Furthermore, dietary oleamide suppressed the mRNA expression of inflammation-related factors in the abdominal fat of mice housed in small cages. Hence, these results indicate that although housing mice in small cages induces obesity and increases abdominal fat mass, dietary oleamide suppresses the obesity.

Oleamide as analyte protectant in GC analysis of THC and its metabolites in blood.

Methods for the analysis of cannabinoids in bio-matrices are continually being developed, to achieve a proper sensitivity required for their detection and accuracy in their quantification. The presented paper shows that the analytical sensitivity of GC-MS to THC and its metabolites in blood samples can be significantly increase by oleamide (OLA) addition to the examined sample, which evokes the matrix effect of transient character. The magnitude of signal increment resulting from oleamide presence in the examined sample is the greatest for THC metabolites and depends on oleamide concentration in the examined sample. The use of transient matrix effect to increase the sensitivity of the analysis can be applied not only in QuEChERS procedure, which is applied in the described experiments, but also in other blood sample preparation methods. Evoking the transient matrix effect by means of OLA in the experimental analytical quantitation of THC and its metabolites in blood allowed to lower limit of detection (LOD) approximately by 20.5%, 87.6% and 90.1% in the case of THC, THC-OH and THC-COOH, respectively.

Assessment of NSAIDs as potential inhibitors of the fatty acid amide hydrolase I (FAAH-1) using three different primary fatty acid amide substrates in vitro.

BACKGROUND: Pain relief remains a major subject of inadequately met need of patients. Therapeutic agents designed to treat pain and inflammation so far have low to moderate efficiencies with significant untoward side effects. FAAH-1 has been proposed as a promising target for the discovery of drugs to treat pain and inflammation without significant adverse effects. FAAH-1 is the primary enzyme accountable for the degradation of AEA and related fatty acid amides. Studies have revealed that the simultaneous inhibition of COX and FAAH-1 activities produce greater pharmacological efficiency with significantly lowered toxicity and ulcerogenic activity. Recently, the metabolism of endocannabinoids by COX-2 was suggested to be differentially regulated by NSAIDs. METHODS: We analysed the affinity of oleamide, arachidonamide and stearoylamide at the FAAH-1 in vitro and investigated the potency of selected NSAIDs on the hydrolysis of endocannabinoid-like molecules (oleamide, arachidonamide and stearoylamide) by FAAH-1 from rat liver. NSAIDs were initially screened at 500 µM after which those that exhibited greater potency were further analysed over a range of inhibitor concentrations. RESULTS: The substrate affinity of FAAH-1 obtained, increased in a rank order of oleamide < arachidonamide < stearoylamide with resultant Vmax values in a rank order of arachidonamide > oleamide > stearoylamide. The selected NSAIDs caused a concentration-dependent inhibition of FAAH-1 activity with sulindac, carprofen and meclofenamate exhibiting the greatest potency. Michaelis-Menten analysis suggested the mode of inhibition of FAAH-1 hydrolysis of both oleamide and arachidonamide by meclofenamate and indomethacin to be non-competitive in nature. CONCLUSION: Our data therefore suggest potential for study of these compounds as combined FAAH-1-COX inhibitors.

Neonatal Supplementation of Oleamide During Suckling Promotes Learning Ability and Memory in Adolescent Mice.

BACKGROUND: Fatty acid amides (FAMs) are present in breast milk. Oleamide (ODA), a member of the FAM family, has been reported to affect learning and memory-related abilities in animal experiments. OBJECTIVES: This study aimed to characterize the temporal changes of FAMs in human milk and sought to examine the effect of ODA supplementation during suckling on postweaning cognitive performance in mice. METHODS: FAMs were measured in human milk (postpartum 1-24 wk) by ultra-performance liquid chromatography-triple quadruple mass spectrometry (UPLC-TQ-MS) analysis. We supplemented neonatal C57BL/6J mice of both sexes with vehicle (control), 5 mg/(kg · day) ODA (L-ODA), or 25 mg/(kg · day) ODA (H-ODA) throughout suckling by oral gavage. After weaning, the Morris water maze test and novel object recognition test were performed. Neurogenesis, spinal morphogenesis in the dentate gyrus (DG) region, and hippocampal expression of synaptic markers were analyzed. Data were analyzed by ANOVA and repeated-measures ANOVA. RESULTS: ODA (0.566-1.31 mg/L) was the most abundant FAM in breast milk, followed by palmitamide (0.135-0.269 mg/L) and linoleamide (0.046-0.242 mg/L). Compared with the control group, the H-ODA group demonstrated shorter escape latency, shorter travel distance, 113% more platform crossing, and 48% greater discrimination index in behavioral tests (P < 0.05). Additionally, the H-ODA group showed a higher density of 5-ethynyl-2'-deoxyuridine (EdU)+ and EdU+& doublecortin (DCX)+ cells (62% and 53%, respectively), and 52% greater spine density in the DG region than the control group (P < 0.05). The synaptic markers, postsynaptic density protein 95 (PSD95) and synaptophysin (SYP), were upregulated in the H-ODA group compared with the control group (P < 0.05). The L-ODA group also showed shorter escape latency in behavioral tests and 27% greater spine density in the DG region than the control group (P < 0.05). CONCLUSIONS: ODA is the most common FAM in human milk. ODA supplementation during suckling promotes learning and memory-related abilities in adolescent mice by augmenting hippocampal neuronal proliferation and boosting synaptic plasticity.

Oleamide, a Sleep-Inducing Supplement, Upregulates Doublecortin in Hippocampal Progenitor Cells via PPARα.

BACKGROUND: Doublecortin (DCX), a microtubule associated protein, has emerged as a central biomarker of hippocampal neurogenesis. However, molecular mechanisms by which DCX is regulated are poorly understood. OBJECTIVE: Since sleep is involved with the acquisition of memory and oleamide or 9-Octadecenamide (OCT) is a sleep-inducing supplement in human, we examined whether OCT could upregulate DCX in hippocampal progenitor cells (HPCs). METHODS: We employed real-time PCR, western blot, immunostaining, chromatin immunoprecipitation, lentiviral transduction in HPCs, and the calcium influx assay. RESULTS: OCT directly upregulated the transcription of Dcx in HPCs via activation of peroxisome proliferator-activated receptor α (PPARα), a lipid-lowering transcription factor. We observed that, HPCs of Ppara-null mice displayed significant impairment in DCX expression and neuronal differentiation as compared to that of wild-type mice. Interestingly, treatment with OCT stimulated the differentiation process of HPCs in wild-type, but not Ppara-null mice. Reconstruction of PPARα in mouse Ppara-null HPCs restored the expression of DCX, which was further stimulated with OCT treatment. In contrast, a dominant-negative mutant of PPARα significantly attenuated the stimulatory effect of OCT on DCX expression and suppressed neuronal differentiation of human neural progenitor cells. Furthermore, RNA microarray, STRING, chromatin immunoprecipitation, site-directed mutagenesis, and promoter reporter assay have identified DCX as a new target of PPARα. CONCLUSION: These results indicate that OCT, a sleep supplement, directly controls the expression of DCX and suggest that OCT may be repurposed for stimulating the hippocampal neurogenesis.

Forensic Analysis of Polymeric Carpet Fibers Using Direct Analysis in Real Time Coupled to an AccuTOF™ Mass Spectrometer.

Polymeric fibers are encountered in numerous forensic circumstances. This study focused on polymeric carpet fibers most encountered at a crime scene, which are nylons, polyesters and olefins. Analysis of the multiple polymer types was done using Direct Analysis in Real Time (DART™) coupled to an Accurate time-of-flight (AccuTOF™) mass spectrometer (MS). A DART gas temperature of 275 °C was determined as optimal. Twelve olefin, polyester, and nylon polymer standards were used for parameter optimization for the carpet fiber analysis. A successful identification and differentiation of all twelve polymer standards was completed using the DART-AccuTOF™. Thirty-two carpet samples of both known and unknown fiber composition were collected and subsequently analyzed. All samples with known fiber compositions were correctly identified by class. All of the remaining carpet samples with no known composition information were correctly identified by confirmation using Fourier-transform infrared spectroscopy (FTIR). The method was also capable of identifying sub-classes of nylon carpet fibers. The results exhibit the capability of DART-AccuTOF™ being applied as an addition to the sequence of tests conducted to analyze carpet fibers in a forensic laboratory.

Improving the sensitivity of estimating CBD and other xenobiotics in plasma samples: Oleamide-induced transient matrix effect.

The paper discusses the matrix effect evoked by oleamide (OLA), a compound frequently found in samples processed and/or stored in lab polypropylene vials or disposable syringes. In the case of many substances a higher response for their samples containing OLA than for net solutions is observed. The analyte signal gain resulting from OLA presence in the examined sample depends on the ratio of OLA concentration to analyte concentration. A characteristic feature of the matrix effect evoked by oleamide is its short duration, which makes the chromatographic data (retention value and signal magnitude of examined compound) repeatable and reproducible. The identified "transient matrix effect" may significantly increase the sensitivity of many analytical procedures employing GC. Evoking the transient matrix effect by means of OLA in the experimental analytical quantitation of cannabidiol in plasma allowed to lower its limit of detection (LOD) by more than 50 %.

Quantification of tire tread wear particles in microparticles produced on the road using oleamide as a novel marker.

In general, tire tread rubber compounds contain oleamide for improvement of manufacturing processibility, mold release characterization, and abrasion resistance. Tire tread wear particles (TWPs) are one of major contributors to microplastic emissions. In this study, a novel analytical method for quantification of TWP in microparticles produced on the road (road dust, MPRs) was developed by employing oleamide as a new marker. MPRs were collected at bus stops in autumn, winter, and summer seasons. MPRs of 38-63, 63-106, 106-212, and 212-500 µm obtained by size separation were employed for the analysis. Rubber components for bus and passenger car tire tread compounds were identified using pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS). Oleamide was extracted from the MPRs with acetone and was identified using GC/MS. The oleamide concentration was analyzed using GC equipped with flame ionization detector (FID). The TWP contents of the MPRs were determined using the oleamide concentrations and the reference compound formulations. In order to reduce the sampling errors, each experiment was carried out five times and the results were averaged. The TWP contents of the MPRs were 1.4-4.7 wt% and were different according to the sampling seasons and places. The TWP contents were increased by increasing the traffic volume and the temperature.

Cannabinoids and Sleep/Wake Control.

The sleep-wake cycle is a complex process that includes wake (W), non-rapid-eye-movement (NREM) and rapid-eye-movement (REM) sleep. Each phase is regulated by specialized brain structures that, by means of different neurotransmitters, maintain the constant expression of the sleep-wake cycle. Molecules like orexin, serotonin, noradrenaline, histamine, for waking; GABA, adenosine, prostaglandins, for NREM sleep and acetylcholine and glutamate for REM sleep, among other molecules are responsible for the expression and maintenance of each phase. When the endocannabinoid system was being described for the first time, almost three decades ago, oleamide's sleep promoting properties were highlighted. Nowadays, enough evidence has been cumulated to support the endocannabinoid system role in the sleep-wake cycle regulation. The endocannabinoids oleamide anandamide, and 2-arachidonylglycerol promote NREM and/or REM sleep via the CB1R, thereby making this system a target to treat sleep disorders, such as insomnia.

Oleamide in Ipomoea and Dillenia Species and Inflammatory Activity Investigated through Ion Channel Inhibition.

BACKGROUND: Oleamide is an essential substance for human health. So, the plants with high oleamide content are great sources for health care products. OBJECTIVE: This study is conducted to investigate the quality of oleamide in plants and test the bioactivity in the selected two studied species. METHODS: The three Ipomoea and five Dillenia species including Ipomoea alba, Ipomoea aquatica and Ipomoea pes-caprae, and Dillenia indica, Dillenia obovata, Dillenia ovata, Dillenia parviflora and Dillenia pentagyna were investigated for the quantity of oleamide by high-performance liquid chromatography. The biological activity test was conducted on the powder formulation of the chosen plants, Dillenia ovata and Dillenia parviflora at a ratio of 30:70, for anti-inflammatory activity ex vivo on a panel of molecular targets through ion channel inhibition including voltage-gated sodium channel, voltage-gated potassium channel, and the cardiac ion as human ether-a-go-go related gene. RESULTS: The results showed that the leaf extracts of I. aquatica and D. ovata gave the highest and subsequent oleamide quantity i.e. 7.52 and 5.17 mg/g, respectively. Out of the Dillenia formulation which contained various compounds, oleamide showed the highest percentages of inhibition at 8.0-20.0%, and 6.2-14.2% in voltage-gated sodium channel, and voltage-gated potassium channel which had slightly lower values than the oleamide standard, and no effect as 0.0% value inhibition in the cardiac ion channel. CONCLUSION: The Dillenia formulation exhibits anti-inflammatory activity without affecting the heart. Accordingly, the three studied Ipomoea and three studied Dillenia species may be used for the same activity as a single component or formulation with effective solvent for disease treatments.

In Vitro Bioassay-Guided Identification of Anticancer Properties from Moringa oleifera Lam. Leaf against the MDA-MB-231 Cell Line.

Moringa oleifera Lam. (MO) is a medicinal plant distributed across the Middle East, Asia, and Africa. MO has been used in the traditional treatment of various diseases including cancer. This study aimed to perform bioassay-guided fractionation and identification of bioactive compounds from MO leaf against MDA-MB-231 breast cancer cells. MO leaf was sequentially extracted with hexane, ethyl acetate (EtOAc), and ethanol. The most effective extract was subjected to fractionation. MO extract and its derived fractions were continuously screened for anti-cancer activities. The strongest fraction was selected for re-fractionation and identification of bioactive compounds using LC-ESI-QTOF-MS/MS analysis. The best anticancer activities were related to the fraction no. 7-derived crude EtOAc extract. This fraction significantly reduced cell viability and clonogenic growth and increased cells apoptosis. Moreover, sub-fraction no. 7.7-derived fraction no. 7 was selected for the identification of bioactive compounds. There were 10 candidate compounds tentatively identified by LC-ESI-QTOF-MS. Three of identified compounds (7-octenoic acid, oleamide, and 1-phenyl-2-pentanol) showed anticancer activities by inducing cell cycle arrest and triggering apoptosis through suppressed Bcl-2 expression which subsequently promotes activation of caspase 3, indicators for the apoptosis pathway. This study identified 10 candidate compounds that may have potential in the field of anticancer substances.

Oleamide Induces Cell Death in Glioblastoma RG2 Cells by a Cannabinoid Receptor-Independent Mechanism.

The endocannabinoid system has been associated with antiproliferative effects in several types of tumors through cannabinoid receptor-mediated cell death mechanisms. Oleamide (ODA) is a CB1/CB2 agonist associated with cell growth and migration by adhesion and/or ionic signals associated with Gap junctions. Antiproliferative mechanisms related to ODA remain unknown. In this work, we evaluated the effects of ODA on cell viability and morphological changes in a rat RG2 glioblastoma cell line and compared these effects with primary astrocyte cultures from 8-day postnatal rats. RG2 and primary astrocyte cultures were treated with ODA at increasing concentrations (25, 50, 100, and 200 µM) for different periods of time (12, 24, and 48 h). Changes in RG2 cell viability and morphology induced by ODA were assessed by viability/mitochondrial activity test and phase contrast microscopy, respectively. The ratios of necrotic and apoptotic cell death, and cell cycle alterations, were evaluated by flow cytometry. The roles of CB1 and CB2 receptors on ODA-induced changes were explored with specific receptor antagonists. ODA (100 µM) induced somatic damage, detachment of somatic bodies, cytoplasmic polarization, and somatic shrinkage in RG2 cells at 24 and 48 h. In contrast, primary astrocytes treated at the same ODA concentrations exhibited cell aggregation but not cell damage. ODA (100 µM) increased apoptotic cell death and cell arrest in the G1 phase at 24 h in the RG2 line. The effects induced by ODA on cell viability of RG2 cells were independent of CB1 and CB2 receptors or changes in intracellular calcium transient. Results of this novel study suggest that ODA exerts specific antiproliferative effects on RG2 glioblastoma cells through unconventional apoptotic mechanisms not involving canonical signals.

Oleamide, a Bioactive Compound, Unwittingly Introduced into the Human Body through Some Plastic Food/Beverages and Medicine Containers.

The purpose of the study was to investigate the migration of oleamide, a polymer lubricant, and a bioactive compound, from various plastic, marketed containers for food/beverages and medicines into polymer contact liquid. Methanol, food/medicine simulants or real samples were used to extract polymer leachables and extractables. Migrated oleamide into polymer contact liquids was determined by ultra-high performance liquid chromatography coupled to mass spectrometry (UHPLC-MS). The concentration of oleamide in the extracts of medicinal and insulin syringes was 7351 ng mL-1 and 21,984 ng mL-1, respectively. The leachates of intravenous (i.v.) infusion bottle, medicinal and insulin syringes contained 17 ng mL-1, 12 ng mL-1 and 152 ng mL-1, respectively. Oleamide in the extracts of dummies ranged from 30 to 39 ng mL-1, while in the leachates of baby bottles, from 12 to 23 ng mL-1. Leachates of soft drink bottles contained from 6 to 15 ng mL-1 oleamide, milk bottles from 3 to 9 ng mL-1, liquid yogurt bottles 17 ng mL-1 and water bottles from 11 to 18 ng mL-1. Bottled real matrices of oil and milk contained oleamide in the range from 217 to 293 ng mL-1. Moreover, the source of migrated oleamide (e.g., containers, caps, other parts) was identified. Oleamide is listed in the current EU regulations without a specific migration limit. Accordingly, these values are considered of no concern, unless future toxicological studies prove the opposite.