Oral Bioavailability and Metabolism of Hydroxytyrosol from Food Supplements.

Table olives and olive oils are the main dietary sources of hydroxytyrosol (HT), a natural antioxidant compound that has emerged as a potential aid in protection against cardiovascular risk. Bioavailability studies with olive oils showed that HT is bioavailable from its free form and from conjugated forms such as oleuropein and its aglycone. Still, its low dietary intake, poor bioavailability, and high inter-individual variability after absorption through the gastrointestinal tract hamper its full benefits. In a randomized, controlled, blinded, cross-over study, we investigated the impact of HT metabolism and bioavailability by comparing two olive-derived watery supplements containing different doses of HT (30.58 and 61.48 mg of HT/dosage). Additionally, HT-fortified olive oil was used in the control group. To this aim, plasma and urine samples were evaluated in 12 healthy volunteers following the intake of a single dose of the supplements or fortified olive oil. Blood and urine samples were collected at baseline and at 0.5, 1, 1.5, 2, 4, and 12 h after intake. HT and its metabolites were analyzed using UHPLC-DAD-MS/MS. Pharmacokinetic results showed that dietary HT administered through the food supplements is bioavailable and bioavailability increases with the administered dose. After intake, homovanillic acid, HT-3-O-sulphate, and 3,4-dihydroxyphenylacetic acid are the main metabolites found both in plasma and urine. The maximum concentrations in plasma peaked 30 min after intake. As bioavailability of a compound is a fundamental prerequisite for its effect, these results promise a good potential of both food supplements for protection against oxidative stress and the consequent cardiovascular risk.

AJ76 and UH232 as potential agents for diagnosing early-stage Parkinson's disease.

We recently reported that the "Dopamine Neuron Challenge Test" (DNC Test), a diagnostic method that measures the levels of dopamine metabolites in cerebrospinal fluid (CSF) and plasma samples after pharmacologically inducing a transient dopamine release, can detect early-stage Parkinson's disease (PD) with high sensitivity and selectivity in mouse models. The use of haloperidol in the original DNC test to challenge dopamine neurons was less than ideal, as it may cause extrapyramidal motor symptoms. Here we report an improved DNC Test, in which the original challenging agents, haloperidol and methylphenidate, are replaced by a single challenging agent, a dopamine autoreceptor preferring antagonist AJ76 or UH232. We show that the improved DNC Test can achieve the same level of sensitivity and selectivity in detecting early PD in a mouse model without causing motor side effects. These findings significantly improve the practicality of using the DNC Test as a screening or diagnostic test for detecting early-stage PD in the high-risk population in humans.

Stress during puberty exerts sex-specific effects on depressive-like behavior and monoamine neurotransmitters in adolescence and adulthood.

Psychiatric disorders including major depression are twice as prevalent in women compared to men. This sex difference in prevalence only emerges after the onset of puberty, suggesting that puberty may be a sensitive period during which sex-associated vulnerability to stress-related depression might become established. Thus, this study investigated whether stress occurring specifically during the pubertal window of adolescence may be responsible for this sex difference in depression vulnerability. Male and female rats were exposed to a three-day stress protocol during puberty (postnatal days 35-37 in females, 45-47 in males) and underwent behavioral tests in adolescence or adulthood measuring anhedonia, anxiety-like behavior, locomotor activity and antidepressant-like behavior. Brainstem and striatum tissue were collected from a separate cohort of behavioral test-naïve rats in adolescence or adulthood to quantify the effect of pubertal stress on monoamine neurotransmitters. Pubertal stress increased immobility behavior in the forced swim test in both sexes in adolescence and adulthood. In adolescence, pubertal stress altered escape-oriented behaviors in a sex-specific manner: decreasing climbing in males but not females and decreasing swimming in females but not males. Pubertal stress decreased adolescent brainstem noradrenaline specifically in females and had opposing effects in adolescent males and females on brainstem serotonin turnover. Pubertal stress induced anhedonia in the saccharin preference test in adult males but not females, an effect paralleled by a male-specific decrease in striatal dopamine turnover. Pubertal stress did not significantly impact anxiety-like behavior or locomotor activity in any sex at either age. Taken together, these data suggest that although pubertal stress did not preferentially increase female vulnerability to depressive-like behaviors compared to males, stress during puberty exerts sex-specific effects on depressive-like behavior and anhedonia, possibly through discrete neurotransmitter systems.

The microbiota catabolites of quercetin glycosides concertedly enhance the resistance against acetaldehyde-induced oxidative stress.

3,4-Dihydroxyphenylacetic acid (DOPAC) and 3-hydroxyphenylacetic acid (OPAC) are the predominant catabolites of quercetin glycosides, such as quercetin 4'-O-ß-glucoside from the onion, produced by intestinal microbiota. Although each catabolite has been reported to protect the cells from acetaldehyde-induced cytotoxicity, the effect of their combination remains to be clarified. The purpose of this study was to determine whether the combination of DOPAC and OPAC enhances the resistance against the acetaldehyde-induced oxidative stress in the cultured hepatocytes. The pretreatment of the combination of DOPAC (5 µM) and OPAC (5 µM) showed significant protection against the acetaldehyde- and hydrogen peroxide-induced cytotoxicity, even though each compound at the same concentration did not. This combination also significantly inhibited the intracellular dichlorofluorescin diacetate-detectable reactive oxygen species (ROS) level, whereas the solo treatment did slightly, suggesting that reducing mechanisms of ROS or compounds that enhance ROS production are involved in the cytoprotective effect. The combinatory treatment significantly enhanced the gene expression of not only the aldehyde dehydrogenases (ALDHs), but also glutamate-cysteine ligase, catalytic subunit, the first rate-limiting enzyme of glutathione (GSH) synthesis. Accordingly, both the intracellular GSH level and the total ALDH activity were enhanced by DOPAC plus OPAC. Involvement of GSH in the cytoprotection as well as ALDH up-regulation by the combination was confirmed by the experiments using a GSH biosynthesis inhibitor, buthionine sulfoximine. Taken together, the present results suggested that the quercetin microbiota catabolites concertedly protect the cells from acetaldehyde through a pre-enhanced resistance against oxidative stress by the GSH-dependent up-regulation of ALDHs.

3,4-Dihydroxyphenylacetic acid ameliorates gut barrier dysfunction via regulation of MAPK-MLCK pathway in type 2 diabetes mice.

AIMS: This study aims to investigate whether 3,4-dihydroxyphenylacetic acid (DHAA) can improve gut barrier function by inhibiting the mitogen-activated protein kinase (MAPK) - myosin light-chain kinase (MLCK) signaling pathway in type 2 diabetes (T2D) mice. MAIN METHODS: T2D mice were induced by a high-fat diet combined with streptozotocin. T2D mice were intragastrically administered with DHAA at 75 and 150 mg kg/body weight per day for 4 weeks. Blood glucose, insulin level, oxidative stress and inflammatory cytokines were measured. TJ (tight junction) protein and MAPK-MLCK pathway-related proteins were analyzed by western blot. KEY FINDINGS: DHAA alleviated hyperglycemia and decreased insulin resistance of T2D mice. It also decreased oxidative stress via increased glutathione (GSH) and total superoxide dismutase (T-SOD) activities and reduced containing malondialdehyde (MDA). DHAA exhibited a significant anti-inflammatory effect by decreasing the level of pro-inflammatory cytokines lipopolysaccharide (LPS) and interleukin (IL)-6 and increasing that of anti-inflammatory cytokine IL-10. More importantly, DHAA improved gut barrier function by enhancing tight junction protein expression and inhibiting the MAPK-MLCK signaling pathway. SIGNIFICANCE: DHAA could reduce oxidative stress, decrease inflammatory response, and improve intestinal function in T2D mice, which may help to relieve the symptoms of T2D.

Ketogenic therapy for Parkinson's disease: A systematic review and synthesis without meta-analysis of animal and human trials.

OBJECTIVE: The aim of the present systematic review was to assess the efficacy of ketogenic therapy in Parkinson's disease (PD), using all available data from randomized controlled trials (RCTs) on humans and animal studies with PD models. DESIGN: Systematic review of in vivo studies. METHODS: Studies related to the research question were identified through searches in PubMed, Cochrane Central Register of Controlled Trials (CENTRAL), Scopus, clinicaltrials.gov and the gray literature, from inception until November 2021. Rayyan was employed to screen and identify all studies fulfilling the inclusion criteria. Cochrane's revised Risk of Bias 2.0 and SYRCLE tools evaluated bias in RCTs and animal studies, respectively. An effect direction plot was developed to synthesize the evidence of the RCTs. RESULTS: Twelve studies were identified and included in the qualitative synthesis (4 RCTs and 8 animal trials). Interventions included ketogenic diets (KDs), supplementation with medium-chain triglyceride (MCT) oil, caprylic acid administration and ketone ester drinks. The animal research used zebrafish and rodents, and PD was toxin-induced. Based on the available RCTs, ketogenic therapy does not improve motor coordination and functioning, cognitive impairment, anthropometrics, blood lipids and glycemic control, exercise performance or voice disorders in patients with PD. The evidence is scattered and heterogenous, with single trials assessing different outcomes; thus, a synthesis of the evidence cannot be conclusive regarding the efficacy of ketogenic therapy. On the other hand, animal studies tend to demonstrate more promising results, with marked improvements in locomotor activity, dopaminergic activity, redox status, and inflammatory markers. CONCLUSIONS: Although animal studies indicate promising results, research on the effect of ketogenic therapy in PD is still in its infancy, with RCTs conducted on humans being heterogeneous and lacking PD-specific outcomes. More studies are required to recommend or refute the use of ketogenic therapy in PD.

Melanopsin modulates refractive development and myopia.

Myopia, or nearsightedness, is the most common form of refractive abnormality and is characterized by excessive ocular elongation in relation to ocular power. Retinal neurotransmitter signaling, including dopamine, is implicated in myopic ocular growth, but the visual pathways that initiate and sustain myopia remain unclear. Melanopsin-expressing retinal ganglion cells (mRGCs), which detect light, are important for visual function, and have connections with retinal dopamine cells. Here, we investigated how mRGCs influence normal and myopic refractive development using two mutant mouse models: Opn4-/- mice that lack functional melanopsin photopigments and intrinsic mRGC responses but still receive other photoreceptor-mediated input to these cells; and Opn4DTA/DTA mice that lack intrinsic and photoreceptor-mediated mRGC responses due to mRGC cell death. In mice with intact vision or form-deprivation, we measured refractive error, ocular properties including axial length and corneal curvature, and the levels of retinal dopamine and its primary metabolite, L-3,4-dihydroxyphenylalanine (DOPAC). Myopia was measured as a myopic shift, or the difference in refractive error between the form-deprived and contralateral eyes. We found that Opn4-/- mice had altered normal refractive development compared to Opn4+/+ wildtype mice, starting ∼4D more myopic but developing ∼2D greater hyperopia by 16 weeks of age. Consistent with hyperopia at older ages, 16 week-old Opn4-/- mice also had shorter eyes compared to Opn4+/+ mice (3.34 vs 3.42 mm). Opn4DTA/DTA mice, however, were more hyperopic than both Opn4+/+ and Opn4-/- mice across development ending with even shorter axial lengths. Despite these differences, both Opn4-/- and Opn4DTA/DTA mice had ∼2D greater myopic shifts in response to form-deprivation compared to Opn4+/+ mice. Furthermore, when vision was intact, dopamine and DOPAC levels were similar between Opn4-/- and Opn4+/+ mice, but higher in Opn4DTA/DTA mice, which differed with age. However, form-deprivation reduced retinal dopamine and DOAPC by ∼20% in Opn4-/- compared to Opn4+/+ mice but did not affect retinal dopamine and DOPAC in Opn4DTA/DTA mice. Lastly, systemically treating Opn4-/- mice with the dopamine precursor L-DOPA reduced their form-deprivation myopia by half compared to non-treated mice. Collectively our findings show that disruption of retinal melanopsin signaling alters the rate and magnitude of normal refractive development, yields greater susceptibility to form-deprivation myopia, and changes dopamine signaling. Our results suggest that mRGCs participate in the eye's response to myopigenic stimuli, acting partly through dopaminergic mechanisms, and provide a potential therapeutic target underling myopia progression. We conclude that proper mRGC function is necessary for correct refractive development and protection from myopia progression.

Ninjin'yoeito, a traditional Japanese medicine, increases dopamine content in PC12 cells.

Dementia is exacerbated by loss of appetite and amotivation, and recent studies have indicated that ninjin'yoeito improves anorexia and amotivation. Previous studies suggest that ninjin'yoeito inhibits dopamine-metabolizing enzymes and enhances dopamine signaling. However, whether ninjin'yoeito increases dopamine content in living cells remains unclear. Here, PC12 cells were used to examine whether ninjin'yoeito affects the dopamine metabolic pathway. Dopamine content significantly increased 3 h after treatment ninjin'yoeito extract. Concomitantly, the levels of 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid were significantly reduced. The effects of components of ninjin'yoeito on the dopamine metabolic pathway were also assessed. Treatment with onjisaponin B, nobiletin, and schisandrin, and the ingredients of Polygalae Radix, Citri Unshiu Pericarpium, and Schisandrae Fructus increased dopamine content and decreased its metabolite content in the culture media. Our findings suggest that ninjin'yoeito improves anorexia and amotivation by inhibiting metabolic enzyme and increasing the dopamine content in cells.

Metoprine, a histamine N-methyltransferase inhibitor, attenuates methamphetamine-induced hyperlocomotion via activation of histaminergic neurotransmission in mice.

Metoprine increases the content of histamine in brain by inhibiting histamine N-methyltransferase (HMT), a centrally acting histamine degrading enzyme. We present data demonstrating that pretreatment with metoprine attenuates the hyperlocomotive effects of METH in mice using a multi-configuration behavior apparatus designed to monitor four behavioral outcomes [horizontal locomotion, appetitive behavior (food access), and food and water intake]. Metoprine pretreatment itself induced hyperlocomotion in mice challenged with saline during the large part of light phase. The trend was also observed during the following dark phase. This is the first report that metoprine has a long-lasting locomotor stimulating property. Similarly, in a tail suspension test, a single injection of metoprine significantly reduced total time of immobility in mice, consistent with the idea that metoprine possesses motor stimulating properties. Metoprine pretreatment did not affect other aspects of behavior. Metoprine did not affect the appetitive and drinking behavior while exerted an effect on stereotypy. No stereotyped behavior was observed in mice pretreated with vehicle followed by METH, while stereotyped sniffing was observed in mice pretreated with metoprine followed by METH. The metoprine pretreatment attenuated METH-induced hyperlocomotion during the first 2 h of light phase, suggesting that metoprine-induced locomotor stimulating property might be different from that of METH. The hypothalamic content of histamine (but not its brain metabolite) was increased after metoprine or METH administration. Both METH and metoprine reduced dopamine and histamine turnover in the striatum and the nucleus accumbens and the hypothalamus, respectively, and there is a significant metoprine pretreatment x METH challenge interaction in the histamine turnover. It is likely that metoprine may attenuate METH-induced hyperlocomotion via activation of histaminergic neurotransmission. Metoprine also might induce a long-lasting locomotor stimulating effect via a putative mechanism different from that whereby METH induces the locomotor stimulating effect.

Hemin Prevents Increased Glycolysis in Macrophages upon Activation: Protection by Microbiota-Derived Metabolites of Polyphenols.

Meat consumption plays a critical role in the development of several types of cancer. Hemin, a metabolite of myoglobin produced after meat intake, has been demonstrated to be involved in the cancer initiation phase. Macrophages are key components of the innate immunity, which, upon activation, can prevent cancer development by eliminating neoplastic cells. Metabolic reprogramming, characterized by high glycolysis and low oxidative phosphorylation, is critical for macrophage activation. 3,4-dihydroxyphenylacetic acid (3,4DHPAA) and 4-hydroxyphenylacetic acid (4HPAA), both microbiota-derived metabolites of flavonoids, have not been extensively studied although they exert antioxidant properties. The aim of this study was to determine the effect of hemin on the anticancer properties of macrophages and the role of 3,4DHPAA and 4HPAA in metabolic reprogramming and activation of macrophages leading to the elimination of cancer cells. The results showed that hemin inhibited glycolysis, glycolytic, and pentose phosphate pathway (PPP) enzyme activities and hypoxia-inducible factor-1 alpha (HIF-1α) stabilization, which interferes with macrophage activation (evidenced by decreased interferon-γ-inducible protein 10 (IP-10) release) and their ability to eliminate cancer cells (via cytotoxic mediators and phagocytosis). Hemin also reduced the mitochondrial membrane potential (MMP) and mitochondrial mass in macrophages. 3,4DHPAA and 4HPAA, by stimulating glycolysis and PPP, prevented the impairment of the macrophage anticancer activity induced by hemin. In conclusion, 3,4HPAA and 4HPAA administration could represent a promising strategy for preventing the reduction of macrophage activation induced by hemin.

The Microbiota-Derived Metabolite of Quercetin, 3,4-Dihydroxyphenylacetic Acid Prevents Malignant Transformation and Mitochondrial Dysfunction Induced by Hemin in Colon Cancer and Normal Colon Epithelia Cell Lines.

Meat diet plays a pivotal role in colorectal cancer (CRC). Hemin, a metabolite of myoglobin, produced after meat intake, has been involved in CRC initiation. The compound, 3,4-dihydroxyphenylacetic acid (3,4HPAA) is a scarcely studied microbiota-derived metabolite of the flavonoid quercetin (QUE), which exert antioxidant properties. The aim of this study was to determine the protective effect of 3,4HPAA against malignant transformation (increased cell proliferation, decreased apoptosis, DNA oxidative damage and augmented reactive oxidative species (ROS) levels) and mitochondrial dysfunction induced by hemin in normal colon epithelial cells and colon cancer cells. The effect of 3,4HPAA was assessed in comparison to its precursor, QUE and to a known CRC protective agent, sulforaphane (SFN). The results showed that both, tumor and normal cells, exposed to hemin, presented increased cell proliferation, decreased caspase 3 activity and cytochrome c release, as well as augmented production of intracellular and mitochondrial ROS. In addition, hemin decreased the mitochondrial membrane potential (MMP) and the activity of complexes I and II of the electron transport chain. These effects of hemin were prevented by the action of 3,4HPAA. The metabolite showed to be more active than QUE and slightly less active than SFN. In conclusion, 3,4HPAA administration could represent a promising strategy for preventing malignant transformation and mitochondrial dysfunction in colon epithelia induced by hemin.

Metabolic profile and underlying antioxidant improvement of Ziziphi Spinosae Folium by human intestinal bacteria.

Ziziphi Spinosae Folium, the leaf of Ziziphus jujuba Mill. Var. spinosa (Bunge) Hu ex H. F. Chou (LZJS), is currently used as a healthy tea in China. This study evaluated the chemical components and antioxidant activities of LZJS flavonoid (LZJSF) and fermented LZJSF (FLZJSF) using human intestinal bacteria (HIB) through dynamic fermentation. Eighteen flavonoids were simultaneously identified in LZJSF using UHPLC-Q-Orbitrap-MS method, nine of which were targeted for a HIB metabolism study. Seven small phenolic acids were identified in FLZJSF. Not only at chemical level but also at PC12 cell level, FLZJSF samples fermented for 4 and 6 h showed significant positive correlation between their activities and flavonoid aglycones, which were transformed from LZJSF. However, FLZJSF samples (8 h and longer time) mainly contained phenolic acids and indicated weak activities. Thus, LZJSF was found to result in increased antioxidant activity and could be commercially utilized as a novel functional food.

Single dose pharmacokinetics of intravenous 3,4-dihydroxyphenylacetic acid and 3-hydroxyphenylacetic acid in rats.

3,4-Dihydroxyphenylacetic acid (DOPAC) and 3-hydroxyphenylacetic acid (3-HPAA) are intestinal metabolites of the dietary flavonoid quercetin. DOPAC reportedly showed anxiolytic activity after i.p. administration in rats. The fate of these metabolites after consumption, and the pharmacological properties of 3-HPAA in the body are largely unknown. The aim of the current study was to characterize pharmacokinetic properties of DOPAC and 3-HPAA after intravenous bolus application in rats. UHPLC-MS/MS methods for quantification of DOPAC and 3-HPAA levels in lithium heparin Sprague Dawley rat plasma were developed and validated according to international regulatory guidelines. Non-compartmental and compartmental analyses were performed. Pharmacokinetic profiles of DOPAC and 3-HPAA followed a two-compartment body model, with a fast distribution into peripheral tissues (half-lives of 3.27-5.26 min) and rapid elimination from the body (half-lives of 18.4-33.3 min).

Antioxidant properties of two novel lipophilic derivatives of hydroxytyrosol.

Two novel lipophilic derivatives of the natural olive oil phenol, hydroxytyrosol (HT), were synthesized using 3,4-dihydroxyphenylacetic acid as starting material. Their antioxidant activities and kinetics compared to HT and TBHQ were assessed by Rancimat, Schaal Oven, 2,2-diphenyl-1-picrylhydrazyl (DPPH) and deep-frying methods. All experiments, including kinetic data analysis based on the Arrhenius equation, utilized in assessing antioxidant activity except the DPPH assay revealed that the new lipophilic HT derivatives exhibited much stronger antioxidant activity than hydroxytyrosol. Tert-butylhydroquinone exhibited stronger antioxidant activity in bulk oil at 65 °C than the new HT derivatives, but showed much lower activity at higher temperatures (>110 °C). This demonstrates that the introduction of bulky alkyl moiety to the ortho-diphenolic structure of HT increased its antioxidant activity. It can be concluded that the new lipophilic HT derivatives satisfy industrial demands for bioactive compounds with strong antioxidant potential at high temperatures.

Is there a relationship between dopamine and rhegmatogenous retinal detachment?

Dopamine and its receptors have been widely studied in the neurological conditions and in the retina. In this study, we evaluated the possible role of dopamine in rhegmatogenous retinal detachment (RRD) by comparing the amount of 3,4-dihydroxyphenylacetic acid (DOPAC), a surrogate index of retinal dopamine levels, in the vitreous sample of patients affected by RRD with those affected by macular pucker and vitreous hemorrhage. Our results showed that significantly higher levels of DOPAC were found in the vitreous sample of patients affected by RRD compared with those affected by vitreous hemorrhage and macular pucker (P = 0.002). Specifically, no trace of the substance was found in vitreous hemorrhage and macular pucker samples. A slightly significant positive correlation was found among DOPAC and post-operative best corrected visual acuity (r = 0.470, P = 0.049). No correlation was found between DOPAC and the days elapsed between diagnosis and surgery (P = 0.317). For the first time our findings suggest that DOPAC is released in RRD, but not in other retinal diseases such as vitreous hemorrhage and macular pucker. Moreover, we showed a correlation between visual acuity outcome and the amount of DOPAC in the vitreous. This might have a potential, although still unknown, implication in the pathogenesis of the disease and/or in the associated photoreceptors loss. This study was approved by the Ethics Committee of Rome Tor Vergata University Hospital (R.S.92.10) on September 24, 2010.

A new inbred strain of Fawn-Hooded rats demonstrates mania-like behavioural and monoaminergic abnormalities.

The Fawn-Hooded (FH) rat carries a gene mutation that results in a dysfunctional serotoninergic system. However, previous studies have reported differing features between the FH/Wjd and FH/Har strains. We aimed to compare the behavioural and neurobiological features of FH/HamSlc rats with those of Fischer 344 rats. We performed the open field, elevated minus-maze, Y-maze spontaneous alternation, and forced swim tests to investigate behavioural alterations. We also assessed neurobiological characteristics by quantifying monoamines and their related compounds in the prefrontal cortex, hippocampus, and striatum using high-performance liquid chromatography with an electrochemical detection system. FH/HamSlc rats showed hyperactivity and a high impulsivity tendency in the open field and the elevated minus maze test, but no cognitive dysfunction. In addition, the hyperactivity was suppressed immediately after the forced swim test. FH/HamSlc rats showed low dopamine levels, but high dopamine turnover in the striatum. Serotonin and noradrenaline levels were low in the prefrontal cortex and the hippocampus of FH/HamSlc rats, but high serotonin turnover was observed in the prefrontal cortex, hippocampus, and striatum. FH/HamSlc rats show (1) mania-like behavioural characteristics that are different from those of other strains of FH rats; (2) stimulus dependent suppression of hyperactivity similar to the clinical findings that exercise alleviates the symptoms of bipolar disorder; and (3) monoaminergic dysregulation such as monoamine imbalance and hyperturnover that may be associated with mania-related behavioural characteristics. Thus, the FH/HamSlc rat is a new animal model for mania including bipolar disorder.

Neuroprotective effect of diclofenac on chlorpromazine induced catalepsy in rats.

Neuroinflammation plays a key role in progressive degeneration of dopaminergic cells. Upregulation of prostaglandins and free radicals formation are involved in the mechanisms of cell death in Parkinson's disease (PD). The present study aimed to investigate the neuroprotective effect of diclofenac against chlorpromazine (CPZ) induced catalepsy and motor impairment in mice. Adult Wistar rats treated with CPZ (3 mg/kg/day, IP) were orally dosed with diclofenac and L-dopa/carbidopa for 21 days. Catalepsy was measured after 21 days of dosing by using standard bar test at 30, 60, 90, 120 and 180 min then motor performances were assessed via open field test and wire hanging test. Histopathological investigation and determination of dopamine (DA) and 3,4-Dihydroxyphenylacetic acid (DOPAC) levels of rat's brain was also carried out. We found that CPZ treated group exhibited reduced motor impairment after 21 days of treatment in open field and wire hanging test (P < 0.01) as compared to control group. The cataleptic scores of CPZ treated rats were also significantly increased (P < 0.01) after 21 days of chronic dosing, however diclofenac treated groups showed significant reduction in cataleptic scores with improved motor performances. Histopathology of CPZ treated rats showed marked degeneration with architecture distortion in the mid brain region. Dopaminergic degeneration is confirmed by neurochemical results that showed reduced amount of dopamine and DOPAC levels in mid brain. Moreover, histopathological slides of diclofenac treated rats showed improved architecture with reduced gliosis of mid brain region as well as improved dopamine and DOPAC levels were achieved after 21 days dosing of diclofenac. Taken together, the present work provide an evidence that diclofenac ameliorated behavioral performances by mediating neuroprotection against CPZ induced PD via preventing dopaminergic neuronal cell death.

Efficient biosynthesis of 3, 4-dihydroxyphenylacetic acid in Escherichia coli.

3, 4-Dihydroxyphenylacetic acid (3, 4-DHPA) is a phenolic acid with strong anti-oxidative activity, showing potential applications in food and pharmaceutical industries. In this study, a 3, 4-DHPA biosynthetic pathway was designed by connecting 4-hydroxyphenylacetic acid (4-HPA) biosynthesis with its hydroxylation. The starting strain produced only 46 mg/L of 4-HPA in 48 h. Enhancing the shikimate pathway increased the titer by 19-fold to 923 ± 57 mg/L. Furthermore, pykA and pykF were disrupted to conserve phosphoenolpyruvate for 4-HPA production. With this effort, 4-HPA titer was increased to 1817 ± 55 mg/L. Introducing the hydroxylase HpaBC into the 4-HPA overproducing strain resulted in 3, 4-DHPA production and the best strain produced 1856 ± 67 mg/L of 3, 4-DHPA in shake flask cultures. This work reports de novo biosynthesis of 3, 4-DHPA for the first time and provides a promising alternative for sustainable production of this valuable compound.

Pharmacokinetics of low molecular weight phenolic compounds in gerbil plasma after the consumption of calafate berry (Berberis microphylla) extract.

Calafate is a berry with high concentration of anthocyanins and hydroxycinnamic acids that grows in South Patagonia. To date, no metabolism studies of phenolic compounds using calafate have been carried out. A calafate extract was characterized by HPLC-DAD-ESI-MS/MS. After extract administration (300 mg/kg), a pharmacokinetic study of phenolic compounds in gerbil plasma was performed by GC-MS/MS. Sixteen phenolic acids increased after intake. Phenylacetic acid derivatives exhibit the highest concentration, while main increase of phenolic catabolites was observed 2 h post-intake. 3-hydroxyphenylacetic and phenylacetic acids increased at 4-8 h post-intake. All catabolites found in gerbil plasma exhibit concentration peaks between 0.1 and 1 µM, however no parental anthocyanins were detected. Establish in vivo plasmatic concentration ranges of phenolic compounds derived from polyphenol consumption following WHO recommendations, plays a key role to carry out future in vitro assays in order to correctly assign biological benefits of calafate berry consumption.

Flavonoid metabolism: the interaction of metabolites and gut microbiota.

Several dietary flavonoids exhibit anti-oxidative, anti-inflammatory, and anti-osteoporotic activities relevant to prevention of chronic diseases, including lifestyle-related diseases. Dietary flavonoids (glycoside forms) are enzymatically hydrolyzed and absorbed in the intestine, and are conjugated to their glucuronide/sulfate forms by phase II enzymes in epithelial cells and the liver. The intestinal microbiota plays an important role in the metabolism of flavonoids found in foods. Some specific products of bacterial transformation, such as ring-fission products and reduced metabolites, exhibit enhanced properties. Studies on the metabolism of flavonoids by the intestinal microbiota are crucial for understanding the role of these compounds and their impact on our health. This review focused on the metabolic pathways, bioavailability, and physiological role of flavonoids, especially metabolites of quercetin and isoflavone produced by the intestinal microbiota.