Liquid Chromatography-electrochemical Detection Research Articles

Separation, identification, and fingerprinting of antioxidant components in persimmon (Diospyros kaki) leaves by offline two-dimensional liquid chromatography with electrochemical detection and tandem mass spectrometry.

In this work, the antioxidant components in persimmon (Diospyros kaki) leaves were separated by offline two-dimensional liquid chromatography-electrochemical detection (LC×LC-ECD) and identified by LC-tandem mass spectrometry (LC-MS/MS). A total of 33 antioxidants, mainly proanthocyanidins, and glycosides of kaempferol and quercetin, were identified. The antioxidant assays demonstrated that the fractions collected from the first-dimension LC (1D-LC) possessed considerable radical scavenging capabilities, with correlation coefficients of peak area versus radical scavenging capability of 1,1-diphenyl-2-picrylhydrazyl and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) being 0.9335 and 0.9116, respectively. The fingerprinting showed that 37 peaks were present in all samples. The major antioxidant components of persimmon leaves were the glycosides of kaempferol and quercetin. Finally, fourteen antioxidants were quantitatively assessed. Offline LC×LC provided high peak capacity and separation; ECD enabled specific screening and detection of antioxidant components; and MS/MS provided excellent identification capability. In this study, the combination of the three approaches was utilized to screen for antioxidant components in persimmon leaves, with satisfactory findings. In conclusion, this technique is an effective means for rapid analysis of antioxidant components and quality control of medicinal plants, achieving rapid separation of congeners and facilitating more accurate qualitative and quantitative analyses.

Diversity in Pyracantha fortuneana fruits maturity stages enables discrepancy in the phenolic compounds, antioxidant activity, and tyrosinase inhibitory activity.

Pyracantha fortuneana (P. fortuneana) fruit is a wild fruit that is popular because of its delicious taste and numerous nutrients, and phenolic compounds are considered to be the main bioactive components in P. fortuneana fruits. However, the relationship between phenolic compounds and their antioxidant and tyrosinase (TYR) inhibitory activities during the ripening process is still unclear. The study compared the influence of the five developmental stages on the accumulation of phenolic compounds, antioxidant activity, and TYR inhibitory activity in the fruits of P. fortuneana. The compounds were identified by offline two-dimensional liquid chromatography-electrochemical detection (2D-LC-ECD) combined with liquid chromatography-tandem mass spectrometry, and the main active ingredients were quantified. The results showed that stage II had higher total phenolic and flavonoid content, as well as higher antioxidant and TYR inhibitory activity, but the total anthocyanin content was lowest at this stage. A total of 30 compounds were identified by 2D-LC-ECD. Orthogonal partial least squares discriminant analysis screened out six major potential markers, including phenolic acids, procyanidins, and flavonoids. In addition, it was found that caffeoylquinic acids, procyanidins, and flavonoids were higher in stage II than in stages I, III, IV, and V, whereas anthocyanins accumulated gradually from stages III to V. Therefore, this study suggests that the changes in antioxidant and TYR inhibitory activities of P. fortuneana during the five developmental stages may be due to the transformation of procyanidins, caffeoylquinic acids, and phenolic glycosides into other forms during the fruit maturation process. Practical Application: Differences in chemical constituents, antioxidant, and tyrosinase inhibitory activities in fruit maturity stages of P. fortuneana were elucidated to provide reference for rational harvesting and utilization of the fruits and their bioactive components. These findings are expected to provide a comprehensive assessment of the bioactive profile and guide the food industrial production.

The Development and Full Validation of a Novel Liquid Chromatography Electrochemical Detection Method for Simultaneous Determination of Nine Catecholamines in Rat Brain.

Chemical neurotransmission, managed by neurotransmitters, has a crucial role in brain processes such as fear, memory, learning, and pain, or neuropathology such as schizophrenia, epilepsy, anxiety/depression, and Parkinson's disease. The measurement of these compounds is used to elucidate the disease mechanisms and evaluate the outcomes of therapeutic interventions. However, this can be quite difficult because of various matrix effects and the problems of chromatographic separation of analysts. In the current study; for the first time, an optimized and fully validated high-performance liquid chromatography-electrochemical detection (HPLC-EC) method according to Food and Drug Administration and European Medicines Agency Bioanalytical Validation Guidance was developed for the simultaneous analysis of nine neurotransmitter compounds, namely dopamine, homovanilic acid, vanilmandelic acid, serotonin (SER), 5-hydroxyindole-3-acetic acid, 4-hydroxy-3-methoxyphenylglycol, norepinephrine, 3,4 dihydroxyphenylacetic acid, and 3-methoxytyramine and simultaneously determined in rat brain samples. Separation was achieved with 150 mm x 4.6 mm, 2.6 μm Kinetex F5 (Phenomenex, USA) column isocratically, and analysis was carried out by HPLC equipped with a DECADE II EC detector. The method exhibited good selectivity, and the correlation coefficient values for each analyte's calibration curves were > 0.99. The detection and quantification limits ranged from 0.01 to 0.03 ng/mL and 3.04 to 9.13 ng/mL, respectively. The stability of the analyses and method robustness were also examined in detail in the study, and the obtained results are presented statistically. The developed and fully validated method has been successfully applied to actual rat brain samples, and important results have been obtained. In the rat brain sample analysis, the lowest number of SER and the highest amount of noradrenaline were found.

Reference intervals for deconjugated urine metanephrines by Bhattacharya analysis.

Urine metanephrines are used to screen for phaeochromocytoma or paraganglioma (PPGL). Current reference intervals (RI) derived in healthy individuals are not age or sex-stratified, and lower than in hypertensive patients, leading to high false positive rates. This study aims to determine age and sex-stratified RI from a contingent screening population. Patients with 24-h deconjugated urine metanephrines from 3/6/2010 to 27/8/2022 were included (2936 males, 5285 females), initially by liquid chromatography-electrochemical detection (LC-ECD) then liquid chromatography-tandem mass spectrometry (LC-MS/MS). Bhattacharya analysis was used after log transformation to determine age and sex-stratified RI for metanephrine excretion, normetanephrine excretion, metanephrine/creatinine and normetanephrine/creatinine ratios. Normetanephrine excretion increases with age (RI: males: 18-<30years: <3.4µmol/24h, 30-<40years: <3.7µmol/24h, 40+ years: <5.3µmol/24h; females: 18-<30years: <2.7µmol/24h, 30-<40years: <3.1µmol/24h, 40+ years: <3.7µmol/24h), while metanephrine excretion was consistent across adulthood (RI: males: 18+ years: <1.8µmol/24h; females: 18+ years: <1.2µmol/24h). However, normetanephrine/creatinine and metanephrine/creatinine increase steadily with age after early adulthood, likely due to a decrease in muscle mass, with females having higher normetanephrine/creatinine and metanephrine/creatinine ratios. Age and sex-stratified RI were derived for metanephrine excretion, normetanephrine excretion, metanephrine/creatinine and normetanephrine/creatinine ratios. This is expected to reduce false positives while flagging most PPGL.

Zhi-Zi-Hou-Po decoction alleviates depressive-like behavior and promotes hippocampal neurogenesis in chronic unpredictable mild stress induced mice via activating the BDNF/TrkB/CREB pathway

Ethnopharmacological relevanceZhi-Zi-Hou-Po decoction (ZZHP), a traditional Chinese medicine (TCM) classic recipe, has been extensively applied for the remedy of depression. However, the underlying mechanism of ZZHP hasn't been fully elucidated and it needs to be further clarified. Aim of studyThe aim of the study is to uncover the mechanisms of ZZHP's effect on depression. Materials and methodsC57BL/6 mice were employed to establish Chronic Unpredictable Mild Stress (CUMS) models. Behavioral tests were conducted for evaluating the antidepressant effects of ZZHP. Then, the monoamine neurotransmitters in the hippocampus through High Performance Liquid Chromatography Electrochemical Detection (HPLC-ECD) were utilized to assess the effect of ZZHP on the maintenance of monoamine neurotransmitter homeostasis. Immunofluorescence staining and Golgi staining were detected to analyze the effects of ZZHP on neuroplasticity in the hippocampus. Western Blot (WB) was utilized to examine the effects of ZZHP on BDNF/TrkB/CREB pathways. Finally, behavioral tests, WB and immunofluorescence staining were repeated after TrkB receptor antagonist was added to further confirm the underlying mechanism. ResultsOur results shown that ZZHP attenuated depressive-like symptoms in CUMS mice. Moreover, ZZHP remarkably reversed the reduction and maintained the homeostasis of monoamine neurotransmitters in the hippocampus. Simultaneously, ZZHP protected neuronal synaptic plasticity and promoted hippocampal neurogenesis. Furthermore, ZZHP stimulated the BDNF/TrkB/CREB pathway in the hippocampus. The addition of TrkB receptor antagonist inhibited the antidepressant effects of ZZHP, suggesting that ZZHP could not work without triggering the BDNF/TrkB/CREB pathway. ConclusionThis study demonstrates that ZZHP can alleviate depressive-like behavior and promote hippocampal neurogenesis in CUMS mice via activating the BDNF/TrkB/CREB pathway.

Leukocyte 8-hydroxy-2'-deoxyguanosine as an oxidative stress marker to predict cardiovascular events and death in chronic hemodialysis patients.

Hemodialysis patients have a markedly increased risk of cardiovascular (CV) morbidity and mortality. Oxidative stress plays a pathogenic role in the progression of atherosclerosis and CV disease among chronic hemodialysis patients. The 8-hydroxy-2'-deoxyguanosine (8-OHdG) content in leukocyte deoxyribonucleic acid (DNA) has been shown as a sensitive and well-known biomarker of oxidant-induced DNA damage in chronic hemodialysis patients. We conducted a retrospective cohort study to investigate the association of leukocyte 8-OHdG and CV events and deaths in patients of chronic hemodialysis. In this study, 217 chronic hemodialysis patients were recruited from 2016 to 2021. The 8-OHdG content of leukocyte DNA was measured by a high-performance liquid chromatography electrochemical detection method. Study outcomes were CV events as well as CV and all-cause deaths. The patients were followed until May 2021. The median follow-up period was 34.8 months. At the end of May 2021, 57 first CV events and 89 all-CV events occurred. Among the first and all CV events, 17 (29.8%) and 32 (36.0%) were fatal, respectively. Multivariate Cox regression analysis showed per 1/10 5 dG increment in leukocyte 8-OHdG values increased risk of CV events (adjusted hazard ratio [aHR], 1.19; 95% CI, 1.10-1.41; p < 0.001), CV death (aHR, 1.27; 95% CI, 1.03-1.72; p = 0.034), and all-cause death (aHR, 1.11; 95% CI, 1.01-1.30; p = 0.038). This is the first study to demonstrate that oxidative stress assessed by 8-OHdG levels of leukocyte DNA predicted CV events as well as CV and all-cause deaths among chronic hemodialysis patients.

Loganin improves chronic unpredictable mild stress-induced depressive-like behaviors and neurochemical dysfunction.

Cornus officinalis Sieb. et Zucc., is a valuable herb commonly used in Chinese medicine clinics. Loganin is a major iridoid glycoside obtained from the traditional Chinese herb Corni Fructus. Loganin, which has been shown to improve depression-like behavior in mice exposed to acute stress, is probably a potential antidepressant candidate. Loganin was evaluated for its effect on chronic unpredictable mild stress (CUMS) induced depressive-like mice, and its action mechanisms were explored. ICR mice were subjected to the CUMS stimulation method to induce depression. The therapeutic effect of loganin on depressive-like behavior was evaluated by a series of behavioral tests such as sucrose preference test (SPT), forced swim test (FST), tail suspension test (TST) and open-field test (OFT). In addition, the serum levels of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) were measured using ELISA. The levels of monoamine neurotransmitters were detected by high performance liquid chromatography-electrochemical detection (HPLC-ECD). The levels of brain-derived neurotrophic factor (BDNF) in the hippocampus were measured using western blot analysis. The results showed that CUMS induced depressive-like behaviors in mice, as indicated by behavioral tests. Administration of loganin increased the sucrose preference in SPT, as well as decreased the immobility time in FST and TST. Loganin could also improve food intake, and increased crossing times in the OFT. In mechanism, loganin restored the secretion of monoamine neurotransmitters, ACTH and CORT to normal levels. In addition, loganin elevated the expression of BDNF in the hippocampus. In conclusion, loganin exerts antidepressant-like effects in CUMS model mice through modulating monoamine neurotransmitters, ACTH, CORT and BDNF. Loganin effectively ameliorated depressive-like symptoms in CUMS-exposed mice by increasing 5-hydroxytryptamine (5-HT) and dopamine (DA) levels, alleviating hypothalamic-pituitary-adrenal (HPA) axis dysfunction, and increasing BDNF expression. In conclusion, the findings of the current study extensive evidence for the application of loganin in stress-associated disorders, specifically targeting depression.

Filter blot method: A simple method for measuring 3-nitrotyrosine in proteins of atmospheric particulate matter

Air pollutants, such as nitrogen dioxide (NO2), ozone (O3), and particulate matter (PM), have been epidemiologically reported to contribute to the onset and exacerbation of asthma. We have previously shown that several proteins in atmospheric PM are allergenic in mouse asthma models and that these proteins are nitrated by atmospheric NO2 and O3 in chemical reactions. Based on these results, the amount of 3-nitrotyrosine (3-NT) in atmospheric PM could be an air pollution marker integrating NO2, O3, and PM. We established a method to measure 3-NT by high-performance liquid chromatography electrochemical detection (HPLC-ECD). Although this method is accurate, it requires a filter treatment process, which is time-consuming and costly for an environmental monitoring tool, in which many samples are measured simultaneously. Therefore, in this study, we investigated a simple immunoblotting method in which atmospheric PM proteins were directly transferred to a polyvinylidene fluoride (PVDF) membrane and measured using an anti-3-NT antibody (the filter blot method). The 3-NT value obtained from this method was significantly correlated (r = 0.809, p < 0.001) with that of the HPLC-ECD method, with a detection power of 0.1 μg/mL for tyrosine nitrated bovine serum albumin equivalents. Multiple regression analysis using the filter blot method showed that the amount of 3-NT in atmospheric PM was significantly associated with the published environmental measurements of O3 and PM in the region. Therefore, the filter blot method may be useful for the environmental monitoring of 3-NT in atmospheric PM.

Effects of artificial light with different spectral composition on eye axial growth in juvenile guinea pigs.

The purpose of the study was to investigate the effect of artificial light with different spectral composition and distribution on axial growth in guinea pigs. Three-week-old guinea pigs were randomly assigned to groupsexposed to natural light, low color temperature light-emitting diode (LED) light, two full spectrum artificial lights (E light and Julia light) and blue light filtered light with the same intensity. Axial lengths of guinea pigs'eyes were measured by A-scan ultrasonography prior to the experiment and every 2 weeks during the experiment. After light exposure for 12 weeks, retinal dopamine (DA), dihydroxy-phenylacetic acid (DOPAC) levelsand DOPAC/DA ratio were analyzed by high-pressure liquid chromatography electrochemical detection and retinal histological structure was observed. Retinal melanopsin expression was detected using Western blot andimmunohistochemistry. After exposed to different kinds of light with different spectrum for 4 weeks, the axial lengths of guinea pigs' eyes in LED group and Julia light group were significantly longer than those of naturallight group. After 6 weeks, the axial lengths in LED light group were significantly longer than those of E light group and blue light filtered group. The difference between axial lengths in E light group and Julia light groupshowed statistical significance after 8 weeks (p<0.05). After 12 weeks of light exposure, the comparison of retinal DOPAC/DA ratio and melanopsin expression in each group was consistent with that of axial length. Inguinea pigs, continuous full spectrum artificial light with no peak or valley can inhibit axial elongation via retinal dopaminergic and melanopsin system.

Profiling of phenolic compounds in domestic and imported extra virgin olive oils in China by high performance liquid chromatography-electrochemical detection

In this study, a high performance liquid chromatography-electrochemical detection (HPLC-ECD) method has been developed for the sensitive determination of 22 phenolic compounds in extra virgin olive oil (EVOO). Chromatographic separation was performed on an Eclipse XDB-C18 column with a gradient methanol/aqueous phosphoric acid elution, and the ECD potential was set at 1.0 V in oxidative mode. Under the optimal conditions, good linearity was obtained for 22 phenolic compounds with coefficients of determination (R2) higher than 0.9971 in the range of 0.005–20.0 μg/mL. The limits of detection (LODs, S/N = 3) for 22 phenolic compounds were in the range of 0.4–6.7 μg/kg by ECD. Satisfactory recoveries ranging from 80.1% to 109.3% were obtained for spiked EVOO samples with relative standard deviations (RSDs) less than 5.4%. The proposed HPLC-ECD method featuring excellent sensitivity and selectivity has been applied to the quantification of phenolic compounds in 61 EVOO samples from seven countries. Good discrimination was achieved between domestic and imported EVOOs based on multivariate statistical approach and their phenolic profiles. These results provide promising perspectives for the use of HPLC-ECD combined with chemometrics for the origin identification of EVOOs.

Reversal of temperature responses to methylone mediated through bi-directional fecal microbiota transplantation between hyperthermic tolerant and naïve rats

ABSTRACT The synthetic cathinone (“bath salt”) methylone induces a hyperthermia response and with chronic administration tolerance to this hyperthermia has been reported. The microbiome-gut-brain axis has been implicated in multiple bodily systems and pathologies, and intentional manipulation of the gut-microbiome has yielded clinically significant results. Here, we examined the effects of bi-directional Fecal Microbiota Transplantation (FMT) between methylone-induced hyperthermic tolerant (MHT) and methylone-naïve (MN) rats. Rats treated with methylone once per week developed tolerance to methylone-induced hyperthermia by the fourth week. Once tolerant, daily bi-directional FMT between the two groups were performed for seven days prior to the next methylone treatment. The FMT abated the developed tolerance in the MHT group. When treated with methylone for the first time following FMT, recipient MN rats displayed significant tolerance to hyperthermia despite it being their initial drug treatment. Post-FMT, MHT rats displayed elevations in norepinephrine and expression of UCP1, UCP3 and TGR5 in brown adipose tissue, with reductions in expression of TGR5 and UCP3 in skeletal muscle. The pre- and post-FMT methylone tolerance phenotypes of transplant recipients are concurrent with changes in the relative abundance of several classes of Proteobacteria, most evident for Gammaproteobacteria and Alphaproteobacteria. MHT recipients demonstrated a marked increase in the relative proportion of the Firmicutes class Erysipelotrichia. These findings suggest that transplantation of gut-microbiomes can confer phenotypic responses to a drug and that the microbiome may be playing a major role in sympathomimetic-mediated hyperthermia. Abbreviations: 3,4-methylenedioxymethamphetamine (MDMA); methylone-induced hyperthermic tolerant (MHT); methylone-naïve (MN); fecal microbiota transplantation (FMT); uncoupling protein (UCP); subcutaneous (sc); intraperitoneal (ip); brown adipose tissue (BAT); skeletal muscle (SKM); sympathetic nervous system (SNS); norepinephrine (NE); quantitative PCR (qRT-PCR); quantification cycle (Cq); High Performance Liquid Chromatography-Electrochemical Detection (HPLC-EC); amplicon sequence variants (ASVs); principal coordinates analysis (PCoA); permutational multivariate analysis (PERMANOVA)

Cornuside ameliorates cognitive impairments in scopolamine induced AD mice: Involvement of neurotransmitter and oxidative stress

Ethnopharmacological relevanceCornus officinalis Sieb. et Zucc., traditional Chinese medicine, has been widely used in the treatment of dementia. Cornel iridoid glycosides of Cornus officinalis is therapeutic to Alzheimer's disease (AD), while its pharmacodynamic material basis is not clear. Cornuside, an iridoid glycoside extracted from of Cornus officinalis Sieb. et Zucc, might be a potential anti-AD candidate. Aim of the studyCornuside was evaluated for its effect on scopolamine induced AD mice, and its action mechanisms were explored. Materials and methodsICR mice were administered with 1 mg/kg scopolamine intraperitoneally to induce amnesia. The therapeutic effect of cornuside of cognitive function was evaluated via series of behavioral tests, including Morris water maze test, step-through test and step-down test. In addition, specific enzyme reaction tests were used to detect the content of acetylcholine (ACh) and malondialdehyde (MDA), as well as the activities of acetylcholinesterase (AChE), butyrylcholinesterase (BuChE), choline acetyltransferase (ChAT), superoxide dismutase (SOD), catalase (CAT), monoamine oxidase (MAO) in the brain. The levels of monoamine neurotransmitters were detected by high performance liquid chromatography-electrochemical detection (HPLC-ECD). ResultsCornuside ameliorated the spatial memory impairment in Morris water maze test and cognitive disruption in step-through and step-down test. Furthermore, cornuside improved the level of ACh by reducing the activities of AChE and BuChE, and increasing the activity of ChAT in hippocampus. Cornuside also increased the levels of monoamine neurotransmitters by inhibiting MAO activity in hippocampus and cortex. In addition, cornuside attenuated MDA by enhancing the activities of SOD and CAT in hippocampus and cortex. ConclusionCornuside improved cognitive dysfunction induced by scopolamine in behavioral tests. The mechanisms of cornuside were further investigated from the aspects of neurotransmitters and oxidative stress. Cornuside could inhibit oxidative stress and neurotransmitter hydrolases, increase ACh and monoamine neurotransmitters, which finally contributed to its therapeutic effect on scopolamine induced amnesia.

Bushen Jieyu Tiaochong Formula reduces apoptosis of granulosa cells via the PERK-ATF4-CHOP signaling pathway in a rat model of polycystic ovary syndrome with chronic stress

Ethnopharmacological relevancePolycystic ovary syndrome (PCOS) is a common and complex endocrine disorder that is also an important cause of infertility. Adverse psychological stress can aggravate the occurrence and development of PCOS. Bushen Jieyu Tiaochong Formula (BJTF), a prescription of Traditional Chinese Medicine (TCM), has been used in the treatment of PCOS and shown to be effective in reducing negative emotion. However, the therapeutic mechanism has yet to be clearly elucidated. In the current study, we investigated the potential mechanism of action of BJTF. Aim of the studyTo investigate the role of PERK-ATF4-CHOP signaling in the molecular mechanisms that mediate the effects of BJTF in a rat model of PCOS, with chronic stress induced by letrozole and a chronic unpredictable mild stress (CUMS) paradigm. Materials and methodsIn addition to the normal control group, the PCOS combined with CUMS model rats were randomly assigned to a model group, a Diane-35 (ethinylestradiol 35 μg/cyproterone acetate 2 mg)-treated positive control group, or one of three BJTF-treated groups receiving a low, medium, or high dose. Behavioral testing, including the sucrose preference test and open field test, was conducted, and hematoxylin and eosin (H&E) staining was used to observe changes in the pathological morphology of ovarian tissue. Free testosterone (FT), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels in serum were quantified by enzyme-linked immunosorbent assays (ELISA). The hippocampal levels of norepinephrine (NE), 5-hydroxytryptamine/serotonin (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) were measured using high-performance liquid chromatography–electrochemical detection (HPLC-ECD). Apoptotic granulosa cells were detected using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Furthermore, quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry were used to detect the expression of glucose-regulated protein 78 (GRP78) and CHOP in the ovarian tissues. The expression levels of GRP78, CHOP, PERK, and ATF4 in ovarian tissues were also measured by western blotting. ResultsTreatment with either BJTF or Diane-35 ameliorated the abnormal cystic dilatation of follicles in the model rats and reduced the serum levels of FT and LH, and the LH/FSH ratio. BJTF treatment also attenuated chronic psychological stress-like behavior and regulated the expression and metabolism of cerebral monoamine neurotransmitters. The efficacy of BJTF was greater than that of Diane-35, with the optimal effects observed at the medium dose. BJTF also lowered the apoptotic index of ovarian granulosa cells and downregulated the expression of GRP78, CHOP, and ATF4. Although the expression level of PERK was not significantly altered by BJTF, the mean PERK expression level was the lowest in the medium-dose BJTF group. ConclusionsAdministration of BJTF has the therapeutic potential to promote the homeostasis of the reproductive endocrine environment and to restore follicular development and ovulation, possibly through the inhibition of the PERK-ATF4-CHOP signaling pathway, leading to downregulation of GRP78 expression to further delay ovarian granule cell apoptosis mediated by endoplasmic reticulum stress (ERS). Moreover, BJTF could improve behavioral performance by regulating cerebral monoamine neurotransmitters in this rat model. These findings provide a new perspective for treating PCOS related to psychological stress using TCM.

Evaluation of the phytochemicals and antioxidant activity of Lophatherum gracile Brongn based on chemical fingerprinting by HPLC with electrochemical detection.

A combinative method using high-performance liquid chromatography-electrochemical detection for fingerprinting and quantitative analysis was developed and successfully applied for the quality evaluation of Lophatherum gracile Brongn leaves collected from 21 geographical locations in China. In the fingerprint analysis, 18 common peaks were observed among the 21 samples, and 10 peaks were identified. Simultaneous quantification of the 10 components was conducted to interpret the variations in these compounds among the L. gracile Brongn leaves originating from different geographical locations. The correlation between the chromatograms and the antioxidant activities of the samples was further studied. The results indicated a linear correlation between the antioxidant activity and the total common peak areas of the fingerprints obtained by high-performance liquid chromatography-electrochemical detection. Importantly, it was found that high-performance liquid chromatography-electrochemical detection fingerprinting can not only determine the quantities of individual components present in such samples but also evaluate the antioxidant activities of the samples. The developed method is a valuable reference for the further study and development of L. gracile Brongn.

Effect of electroacupuncture preconditioning on the contents of dopamine and 5-hydroxytryptamine in lateral hypothalamus area and cerebellar fastigial nucleus of rats with myocardial ischemia-reperfusion injury

To observe the effect of electroacupuncture (EA) preconditioning at heart meridian acupoints on the contents of dopamine (DA) and 5-hydroxytryptamine (5-HT) in lateral hypothalamus area (LHA) and cerebellar fastigial nucleus (FN) in the rats with acute myocardial ischemia-reperfusion injury (MIRI), and explore the role and mechanism of LHA and FN in the effect of EA at heart meridian acupoints against acute MIRI. Sixty SD rats were randomly divided into a sham-operation group, a model group, an EA heart meridian group and an EA lung meridian group, 12 rats in each group, as well as an LHA plus heart meridian group (damage of bilateral LHA) and an FN plus heart meridian group (damage of bilateral FN), 6 rats in each one. Three days after nucleus destruction, EA was applied to "Shenmen" (HT 7) and "Tongli" (HT 5) in the EA heart meridian group, the LHA plus heart meridian group and the FN plus heart meridian group and EA was applied to "Taiyuan" (LU 9) and "Lieque" (LU 7) in the EA lung meridian group, with 1 V in stimulating voltage and 2 Hz in frequency, lasting 20 minutes each time, once a day, for consecutively 7 days before model replication. Except in the sham-operation group, MIRI rat models were duplicated by ligation of the left anterior descending branch of the coronary artery in the rest groups. Using Power lab physiological recorder, ST segment displacement value was recorded before modeling, 30 min after ligation and 120 min after reperfusion separately. The high performance liquid chromatography-electrochemical detection and analysis system was adopted to determine the contents of DA and 5-HT in LHA and FN dialysate after rat modeling in each group. In comparison of ST segment displacement value 30 min after ligation and 120 min after reperfusion among groups, the value in the model group was higher than that in the sham-operation group (P<0.01), those in the EA heart meridian group, the LHA plus heart meridian group, the FN plus heart meridian group and the EA lung meridian group were lower than those in the model group successively (P<0.01) and those in the EA heart meridian group were lower than those in the EA lung meridian group, the LHA plus heart meridian group and the FN plus heart meridian group successively (P<0.01). The contents of DA and 5-HT in FN and LHA in the model group were lower than those in the sham-operation group (P<0.01). Except in the groups with nucleus lesions, the contents of DA and 5-HT in FN and LHA of each intervention group were higher than those in the model group (P<0.01), the contents of DA and 5-HT in FN in the EA heart meridian group were higher than those in the EA lung meridian group and the LHA plus heart meridian group (P<0.01) and the content of 5-HT in LHA was higher than those in the EA lung meridian group and the FN plus heart meridian group (P<0.01) separately, the content of DA in LHA was higher than that in the EA lung meridian group (P<0.01). EA preconditioning at heart meridian acupoints can effectively alleviate myocardial injury in acute MIRI rats, during which, DA and 5-HT in LHA and FN may be the important material basis.

Brain Concentrations of Methylone and Its Metabolites after Systemic Methylone Administration: Relationship to Pharmacodynamic Effects.

3,4-Methylenedioxy-N-methylcathinone (methylone) is a new psychoactive substance with stimulant properties and potential for abuse. Despite its popularity, limited studies have examined relationships between brain concentrations of methylone, its metabolites, and pharmacodynamic effects. The goal of the present study was 2-fold: 1) to determine pharmacokinetics of methylone and its major metabolites-4-hydroxy-3-methoxy-N-methylcathinone (HMMC), 3,4-dihydroxy-N-methylcathinone (HHMC), and 3,4-methylenedioxycathinone (MDC)-in rat brain and plasma and 2) to relate brain pharmacokinetic parameters to pharmacodynamic effects including locomotor behavior and postmortem neurochemistry. Male Sprague-Dawley rats received subcutaneous methylone (6, 12, or 24 mg/kg) or saline vehicle (n = 16/dose), and subgroups were decapitated after 40 or 120 minutes. Plasma and prefrontal cortex were analyzed for concentrations of methylone and its metabolites by liquid chromatography-tandem mass spectrometry. Frontal cortex and dorsal striatum were analyzed for dopamine, 5-HT, and their metabolites by high-performance liquid chromatography-electrochemical detection. Brain and plasma concentrations of methylone and its metabolites rose with increasing methylone dose, but brain methylone and MDC concentrations were greater than dose-proportional. Brain-to-plasma ratios for methylone and MDC were ≥ 3 (range 3-12), whereas those for HHMC and HMMC were ≤ 0.2 (range 0.01-0.2). Locomotor activity score was positively correlated with brain methylone and MDC, whereas cortical 5-HT was negatively correlated with these analytes at 120 minutes. Our findings show that brain concentrations of methylone and MDC display nonlinear accumulation. Behavioral and neurochemical effects of systemically administered methylone are related to brain concentrations of methylone and MDC but not its hydroxylated metabolites, which do not effectively penetrate into the brain. SIGNIFICANCE STATEMENT: Behavioral and neurochemical effects of methylone are related to brain concentrations of methylone and its metabolite MDC but not its hydroxylated metabolites, 4-hydroxy-3-methoxy-N-methylcathinone and 3,4-dihydroxy-N-methylcathinone, which do not effectively penetrate into the brain. Methylone and MDC display nonlinear accumulation in the brain, which could cause untoward effects on serotonin neurons in vulnerable brain regions, including the frontal cortex.

The identification of tyrosine-nitrated human serum albumin in airborne particulate matter from Japan

Atomospheric particulate matter (PM) has been accepted as a risk to human health worldwide. PM contains a variety of components, but it is not clear which components are harmful to human health. For example, there have been many reports on metals, but there have been only a few studies on proteins. Based on an analysis of serum antibodies in a mouse model of PM-induced asthma reported previously, we have been conducting research focusing on the proteins contained in PM. In the present study, we found for the first time that human serum albumin (HSA) is bound to PM. HSA was detected in total suspended particulate matter (TSP) and PM2.5, from various regions in Japan, during all seasons. The presence of HSA in PM was confirmed by proteomics analysis, amino acid analysis, immunoblotting and BD140, which emits fluorescence when bound to a specific HSA site. Moreover, we discovered that TSP-binding HSA was tyrosine-nitrated, one of a chemical modification via environmental atmospheric pollutants using high performance liquid chromatography-electrochemical detection. It was concluded that although the origin of HSA in PM was unclear, HSA is present in the airborne PM of Japan, partially modified for tyrosine nitration.

Exercise preconditioning ameliorates cognitive impairment and anxiety-like behavior via regulation of dopamine in ischemia rats

Cognitive impairment and anxiety are common health problems in acute ischemic stroke patients. Meanwhile, dopamine in the striatal brain region is significantly increased during the acute phase of cerebral ischemia. Besides, the studies shown that striatum and change of striatal dopamine are associated with learning and memory and anxiety. Further, physical exercise has been shown to improve neurocognitive and emotional function in animal models and patients with cerebral ischemia. However, the exact mechanism underlying this effect is unclear. The purpose of this research is to explore the effect of pre-ischemic voluntary wheel running on levels of striatal dopamine, cognition and anxiety in cerebral ischemia rats. Methods: 48 adult male Sprague-Dawley rats were enrolled in this study and divided randomly in following 6 groups: sham group (S group, n = 8), ischemia group (I group, n = 8), 1 week wheel running group (1R group), 4 weeks wheel running group (4R group), 1 week pre-ischemia wheel running group (1RI group, n = 8) and 4 weeks pre-ischemia wheel running group (4RI group, n = 8). After training, cerebral ischemia was induced by permanent bilateral common carotid artery ligation (2-VO) operation. Microdialysis was used to collect dialysates from the striatum immediately from 30 min before ischemia to 90 min after ischemia. High-performance liquid chromatography-electrochemical detection system (HPLC) was used to determine the content of dopamine in the dialysates. Passive avoidance and elevated plus maze test were used to test neurocognitive function 24 h after 2-VO cerebral ischemia. Results: As compare with the constant striatal dopamine level of S group, the striatal dopamine level in I group after ischemia showed a trend of rapid increasing and reached maximum value at the 20 min (P<0.001), then decreased gradually. The striatal dopamine level in 1RI and 4RI group showed the trend were similar to I group, but the increasing magnitude was attenuated. A comparison of the basal striatal dopamine level in 4 groups found that the basal dopamine level in 1RI and 4RI group were higher than S and I group (P<0.001). In passive avoidance task, the retention latency of I group was significantly shorter than S group (P<0.001), and the retention latency of the 1RI, 1R and 4R, 4RI group were longer than I group (P<0.001), there was no significant difference in S, 1RI, 1R, 4R and 4RI group (P>0.05). In elevated plus maze test, the time and entrance numbers of open arms in I group were significantly less than S group (P<0.05), but these indices were no significant difference in S, 1RI, 1R, 4RI and 4RI group. Conclusion: According to our results, 1 or 4 weeks pre-ischemia wheel running can significantly increase the basal dopamine level, attenuate the increase of striatal dopamine induced by cerebral ischemia and improve neurocognitive function in ischemia rats.

Jiaotai Pill () Alleviates Insomnia through Regulating Monoamine and Organic Cation Transporters in Rats.

To reveal the effect and mechanism of Jiaotai Pill (, JTP) on insomniac rats. The insomniac model was established by intraperitoneal injection of p-chlorophenylalanine (PCPA). In behavioral experiments, rats were divided into control, insomniac model, JTP [3.3 g/(kg•d)], and diazepam [4 mg/(kg•d)] groups. The treatment effect of JTP was evaluated by weight measurement (increasement of body weight), open field test (number of crossings) and forced swimming test (immobility time). A high performance liquid chromatography-electrochemical detection (HPLC-ECD) method was built to determine the concentration of monoamine transmitters in hypothalamus and peripheral organs from normal, model, JTP, citalopram [30 mg/(kg•d)], maprotiline [40 mg/(kg•d)] and bupropion [40 mg/(kg•d)] groups. Expressions of serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET) were analyzed by quantitative polymerase chain reaction (qPCR) and Western blot in normal, model and JTP groups. A high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS/MS) method was established to determine the pharmacokinetics, urine cumulative excretion of metformin in vivo, and tissue slice uptake in vitro, which were applied to assess the activity of organic cation transporters (OCTs) in hypothalamus and peripheral organs. Compared with the insomniac model group, the body weight and spontaneous locomotor were increased, and the immobility time was decreased after treatment with JTP (P<0.01). Both serotonin and dopamine contents in hypothalamus and peripheral organs were increased (P<0.01). The norepinephrine content was increased in peripheral organs and decreased in hypothalamus (P<0.05 or P<0.01). At the same time, SERT, DAT, OCT1, OCT2, and OCT3 were down-regulated in hypothalamus and peripheral organs (P<0.05). NET was down-regulated in peripheral organs and up-regulated in hypothalamus (P<0.05 or P<0.01). Moreover, the activity of OCTs in hypothalamus and peripheral organs was inhibited (P<0.05). JTP alleviates insomnia through regulation of monoaminergic system and OCTs in hypothalamus and peripheral organs.

Chronic administration of synthetic cannabidiol induces antidepressant effects involving modulation of serotonin and noradrenaline levels in the hippocampus

Cannabidiol (CBD) is a non-psychotomimetic compound derived from Cannabis sativa. Preclinical and clinical studies have shown therapeutic potential of CBD in a variety of disorders. Despite several research efforts on CBD, its antidepressant activity has been poorly investigated and the exact mechanism of action remains unclear. Thus, this study aimed to further explore the mechanism of CBD after chronic administration (7 days). First, the dose level of CBD that is enough to produce antidepressant effects after chronic administration was explored. Second, the changes in key proteins and neurotransmitters through such methods as real-time polymerase chain reaction (RT-PCR), western blotting, and high-performance liquid chromatography-electrochemical detection (HPLC-ECD) were critically studied. Furthermore, correlation between behavioral phenotypes with protein and neurotransmitters was established and the possible mechanism was herein postulated. The results showed that only the high dose CBD 100 mg/kg chronic administration induced antidepressant-like effects in mice subjected to forced swim test. Chronic CBD 100 mg/kg administration resulted in significant increases in serotonin (5-HT) and noradrenaline (NA) levels in the hippocampus (HPC). Similarly, the chronic administration of CBD 30 mg/kg and CBD 100 mg/kg significantly decreased nuclear factor kappa B (NF-κB) expression in the HPC. Moreover, none of the treatments were observed to induce locomotor effects. Thus, we concluded that chronic administration of CBD (100 mg/kg) induced antidepressant-like effects by increasing 5-HT and NA levels in the HPC. These results shed new light on further discovery of the antidepressant effect of CBD.