Impact of homocysteine on acute ischemic stroke severity: possible role of aminothiols redox status.

BACKGROUND: Acute ischemic stroke (AIS) is one of the most common cerebrovascular diseases which accompanied by a disruption of aminothiols homeostasis. To explore the relationship of aminothiols with neurologic impairment severity, we investigated four aminothiols, homocysteine (Hcy), cysteine (Cys), cysteinylglycine (CG) and glutathione (GSH) in plasma and its influence on ischemic stroke severity in AIS patients. METHODS: A total of 150 clinical samples from AIS patients were selected for our study. The concentrations of free reduced Hcy (Hcy), own oxidized Hcy (HHcy), free reduced Cys (Cys), own oxidized Cys (cysteine, Cyss), free reduced CG (CG) and free reduced GSH (GSH) were measured by our previously developed hollow fiber centrifugal ultrafiltration (HFCF-UF) method coupled with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The concentration ratio of Hcy to HHcy (Hcy/HHcy), Cys to Cyss (Cys/Cyss) were also calculated. The neurologic impairment severity of AIS was evaluated using National Institutes of Health Stroke Scale (NIHSS). The Spearman correlation coefficient and logistic regression analysis was used to estimate and perform the correlation between Hcy, HHcy, Cys, Cyss, CG, GSH, Hcy/HHcy, Cys/Cyss and total Hcy with NIHSS score. RESULTS: The reduced Hcy and Hcy/HHcy was both negatively correlated with NIHSS score in AIS patients with P = 0.008, r=-0.215 and P = 0.002, r=-0.249, respectively. There was no significant correlation of Cys, CG, GSH, HHcy, Cyss, Cys/Cyss and total Hcy with NIHSS score in AIS patients with P value > 0.05. CONCLUSIONS: The reduced Hcy and Hcy/HHcy, not total Hcy concentration should be used to evaluate neurologic impairment severity of AIS patient.

A more accurate indicator to evaluate oxidative stress in rat plasma with osteoporosis.

Background: oxidative stress is linked to various human diseases which developed into the idea of "disrupted redox signaling". Osteoporosis (OP) is a chronic skeletal disorder characterized by low bone mineral density and deterioration of bone microarchitecture among which estrogen deficiency is the main cause. Lack of estrogen leads to the imbalance between oxidation and anti-oxidation in patients, and oxidative stress is an important link in the pathogenesis of OP. The ratio of the reduced to the oxidized thiols can characterize the redox status. However, few methods have been reported for the simultaneous determination of reduced forms and their oxidized forms of thiols in plasma. Methods: we developed a hollow fiber centrifugal ultrafiltration (HFCF-UF) method for sample preparation and validated a high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method to determine two reduced forms of thiols-homocysteine (Hcy), cysteine (Cys) levels and their respective oxidized compounds, homocystine (HHcy) and cystine (Cyss) in rat plasma simultaneously for the first time. Thirty-six female rats were randomly divided into three groups: normal control (NC), oxidative stress (ovariectomy, OVX) and ovariectomy with hydrogen-rich saline administration (OVX + HRS). Results: the validation parameters for the methodological results were within the acceptance criteria. There were both significant differences of Hcy/HHcy (Hcy reduced/oxidized) and Cys/Cyss (Cys reduced/oxidized) in rat plasma between three groups with both p < 0.05 and meanwhile, the p values of malondialdehyde, superoxide dismutase and glutathione peroxidase were all less than 0.01. The value of both Hcy/HHcy and Cys/Cyss were significantly decreased with the change of Micro-CT scan result of femoral neck in OVX group (both the trabecular thickness and trabecular number significantly decreased with a significant increase of trabecular separation) which demonstrate OP occurs. The change of Hcy/HHcy is more obvious and prominent than Cys/Cyss. Conclusions: the Hcy/HHcy and Cys/Cyss could be suitable biomarkers for oxidative stress and especially Hcy/HHcy is more sensitive. The developed method is simple and accurate. It can be easily applied in clinical research to further evaluate the oxidative stress indicator for disease risk factors.

Analysis of free concentrations of lamotrigine and active oxcarbazepine metabolite in clinical patients by hollow-fiber centrifugal ultrafiltration.

Aim: To establish a simple and accurate method to explore the correlation between free and total concentrations of lamotrigine (LTG) and the active oxcarbazepine metabolite monohydroxy derivative (MHD) (10,11-dihydro-10-hydroxycarbamazepine) in clinical patients. Materials & methods: Serum samples were prepared by hollow-fiber centrifugal ultrafiltration and then injected into UPLC for analysis. Results: Absolute recovery was as high as approximately 90.1-98.6% with excellent precision (relative standard deviation <6.7%). Analysis time was reduced to 5 min. There were significant individual differences in the protein binding rates of both LTG and MHD that were probably due to the use of different clinical patients. Conclusion: Free concentrations of LTG and MHD cannot be estimated by total concentration in specific clinical patients. Free drug monitoring of LTG and MHD in clinical therapeutic drug monitoring is important and essential.

Determination of 23 related analytes in bone marrow fluid and hippocampus of C57BL/6 mice following cerebral ischemia-reperfusion injury by HPLC-FLD.

The study of brain diseases has long been of interest to researchers worldwide, and stroke is the third leading cause of death that threatens human health. At the same time, cerebral ischemia-reperfusion injury is closely associated with high rates of disability and mortality. The conditions of the 6-aminoquinolyl N-hydroxysccinimidyl carbamate method for the derivatization of amino acids in the bone marrow fluid and hippocampus of C57BL/6 mice with cerebral ischemia-reperfusion injury were explored and optimized, such as the column temperature, concentration of derivatization reagents and mobile phase concentration. The mobile phase consisted of 20 mm sodium acetate solution (phosphoric acid to adjust pH 5.0) and 60% acetonitrile solution at a flow rate of 1 ml min-1 . The 23 analytes were separated and determined in a gradient elution procedure; the correlation coefficient r was >0.9990 in the range 0.1-8.0 µg ml-1 . The results showed that the content of relevant analytes was significantly changed in the cerebral ischemia-reperfusion injury model, and the method was suitable for the simultaneous determination of 23 amino acids in the bone marrow fluid and hippocampus of C57BL/6 mice.

Contrast-enhanced ultrasound as a valuable imaging modality for characterizing testicular lesions.

Contrast-enhanced ultrasound (CEUS) is a new form of ultrasound (US) that can dynamically display microvessels in a highly sensitive manner. The purpose of this study was to investigate the efficacy of CEUS for characterizing testicular lesions in comparison with conventional US. Forty-seven patients with testicular lesions were enrolled. The histopathology results revealed that 31 cases were neoplastic (11 cases of seminomas, 8 nonseminomatous germ cell tumors, 8 lymphomas, 2 Leydig cell tumors, and 2 nonspecific tumors), and 16 cases were nonneoplastic (8 cases of infarctions, 3 epidermoid cysts, and 5 inflammation). The indicators of shallow lobulated morphology and cystic-solid echogenicity on conventional US were suggestive of germ cell tumors. More indicators on CEUS were found to be useful for characterizing testicular lesions. All the neoplastic lesions showed hyperenhancement on CEUS. Moreover, germ cell tumors presented with heterogeneous enhancement (73.7%, 14/19), a twisted blood vessel pattern, rapid wash-in and wash-out, and peripheral rim hyperenhancement signs. Lymphoma was characterized by nonbranching linear vessel patterns (87.5%, 7/8), rapid wash-in and slow wash-out. In nonneoplastic lesions, infarction and epidermoid cysts showed no enhancement, and abscesses were observed with marginal irregular enhancement. The sensitivity, specificity, and accuracy of CEUS for differentiating between neoplastic and nonneoplastic lesions were 100%, 93.8%, and 97.9%, respectively, and these values were higher than those for conventional US (90.3%, 62.5%, and 80.9%, respectively). CEUS can sensitively reflect the microvascular perfusion in testicular lesions and offers high accuracy for characterizing them.

Establishment of an HPLC Method for Determination of Coumarin-3-Carboxylic Acid Analogues in Rat Plasma and a Preliminary Study on Their Pharmacokinetics.

A simple high performance liquid chromatography (HPLC) method was developed and validated for the determination of coumarin-3-carboxylic acid analogues (C3AA) in rat plasma and a preliminary study on pharmacokinetics. Ferulic acid (FA) was used as the internal standard substance, and coumarin-3-carboxylic acid (C3A) was used as a substitute for quantitative C3AA. After protein precipitation with methanol, the satisfactory separation was achieved on an ODS2 column when the temperature was maintained at 30 ± 2°C. The correlation coefficient r in the C3A linear equation is equal to 0.9990. Pharmacokinetic parameters for t1/2, Tmax, Cmax, area under the curve (AUC)0-t, average residence time (MRT), apparent volume of distribution (V z/F) and clearance (Cl/F) were 1.89 ± 0.03 h, 0.39 ± 0.14 h, 1.81 ± 0.10 g· mL-1 ·h, 7.88 ± 0.24 g·mL-1·h, 3.23 ± 0.14 h, 0.43 ± 0.03 (mg·kg-1)·(g·mL-1)-1·h-1, respectively. The high performance liquid chromatography-photo diode array detector (HPLC-PDA) method established in this study can be used to separate and determine the content of C3AA in plasma of rats after 60% ethanol extraction by gavage. The plasma concentration-time curve and pharmacokinetic parameters reflect the absorption of C3AA in rat blood after oral administration to some extent.

The variation in the levels of 18 amino acids in the cortex and plasma of cerebral ischemia C57BL/6 mice.

Emerging evidence suggests that amino acid (AA) neurotransmitters play important roles in the pathophysiological processes of cerebral ischemia. In this work, an HPLC with fluorescence detection (HPLC-FLR) method was developed for the simultaneous determination of 18 AAs in the cortex and plasma after cerebral ischemia in mice. The ischemia model was prepared by bilateral common carotid artery occlusion, and then the cortex and plasma of the sham, ischemia, and naringenin groups were collected. Based on the protein precipitation method, a simple and effective sample preparation method was developed. The treated sample contained minimal proteins and lipids. The analysis of the sample was performed by the proposed HPLC-FLR method in combination with o-phthalaldehyde. The results showed a statistically significant increase in excitatory AAs (aspartic acid and glutamic acid), inhibitory AAs (glycine and 4-aminobutyric acid), phenylalanine, citrulline, isoleucine, and leucine levels, and a decrease of glutathione and phenylalanine levels when compared with the sham group in the cortex. Besides, the administration of naringenin had significant effects on excitatory AAs, inhibitory AA (glycine), glutamine, tyrosine, phenylalanine, and leucine levels when compared with the sham group in the cortex. These findings could be utilized in studying and clarifying the mechanisms of ischemia.

Simultaneous determination of thirteen substances related to NAFLD in mouse brain tissue using 3-aminobutyric acid as internal standard by HPLC-FLD.

Disorders of certain branched-chain amino acids may be associated with the occurrence and development of non-alcoholic fatty liver disease. Measurement of related branched-chain amino acid levels could provide a reference for the clinical and scientific research of the non-alcoholic fatty liver disease. An established HPLC-FLD method was used to quantify aspartic acid, glutamate, glutamine, glycine, taurine, tyrosine, 4-amino butanoic acid, tryptophan, methionine, valine, phenylalanine, isoleucine and leucine in mouse brain tissue. Brain tissue samples mixed with internal standard (3-aminobutyric acid) were processed, then derivatized with 2-O-phthaldialdehyde, and finally separated on an ODS2 column through gradient elution at a flow rate of 1.0 ml·min-1 . The excitation and emission wavelengths were set at 340 and 455 nm, respectively. The mobile phase A was 100% methanol and the mobile phase B consisted of 30 mmol·L-1 sodium acetate (pH 6.8). The injection volume was 20 µl and the single run time was 45 min. Several parameters, accuracy, precision, and stability, were verified and the results showed the established method had good sensitivity and resolution for all of the 13 compounds and internal standard in mouse brain.

Evaluation on monoamine neurotransmitters changes in depression rats given with sertraline, meloxicam or/and caffeic acid.

Inflammation drives the development of depression and may affect neurotransmitters and thus neurocircuits increase the risk of depression. To investigate the influence of inhibition of inflammatory pathways on the biogenic amine neurotransmitters metabolism in depressive rats, sertraline, and meloxicam, the inhibitors of arachidonic acid - cyclooxygenase-2/lipoxygenase (AA-COX-2/5-LO) pathways, were given to depressive rats. After the development of depression model by chronic unpredictable mild stress (CUMS) for 6 weeks, Successful modeling rats were selected and randomly divided into CUMS group and medication administration group. After given medicine, The biogenic amine neurotransmitters in rat cortex and hippocampus were measured by high-performance liquid chromatography equipped with an electrochemical detector (HPLC-ECD). Compared with the normal group, the concentration of norepinephrine (NE) significantly decreased and the concentrations of Tyrosine (Tyr), Tryptophan (Trp), 3,4-dihydroxyphenyl acetic acid (DOPAC), 3-methoxy-4-hydroxyphenylglycol (MHPG), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) significantly increased in the CUMS group. Sertraline significantly inhibited the elevation of 5-HIAA. Meloxicam inhibited the decrease of NE level in CUMS-induced rat and the increase of Trp, MHPG, and 5-HIAA level in a dose-dependent manner. Caffeic acid inhibited the decrease of NE and the increase of Trp and MHPG in a dose-dependent manner. The inhibition of AA-COX-2/5-LO pathways can improve the behaviors of depression rats and suppress CUMS-induced changes in biogenic amines. Compared with the single-dose lipoxygenase (5-LO) or Cyclooxygenase-2 (COX-2) inhibitor, the combination treatment with meloxicam 1 mg/kg and caffeic acid 10 mg/kg have no significant improvement in CUMS-induced depression behavior and the level of cortical monoamine neurotransmitters and their metabolites.

Simultaneous determination of nine analytes related to the pathogenesis of diabetic encephalopathy in diabetic rat cortex and hippocampus by HPLC-FLD.

The determination of amino acids and monoamine with actions like neurotransmitters or modulators has become increasingly important for studying the relationship between the dysfunction of neurotransmitters and the pathogenesis of diabetic encephalopathy. Here, a high-performance liquid chromatography with fluorescence detection method was developed to simultaneously determine nine monoamines and amino acids including three excitatory neurotransmitters (aspartate, glutamate, and serotonin), four inhibitory neurotransmitters (glycine, γ-aminobutyric acid, taurine, dopamine), a precursor of 5-HT (tryptophan) and methionine using homoserine as the internal standard. The separation was performed on a BDS column with methanol-buffer solution of 35 mmol/L sodium acetate and 5 mmol/L citric acid (pH 6.0) using a simple gradient elution. Several parameters including specificity, precision, and recovery were validated after optimization of the analytical conditions. The developed method was successfully applied to determine the cortex and the hippocampus samples from Sprague-Dawley rats. Our results showed that various neurotransmitters involved in diabetes mellitus may tend to be differentially modulated and present a different alteration tendency at different time course, which might be associated with the duration of diabetes mellitus.

Simultaneous determination of tryptophan, kynurenine, kynurenic acid and two monoamines in rat plasma by HPLC-ECD/DAD.

A high-performance liquid chromatography method with a diode array and an electrochemical detection (HPLC-ECD/DAD) was developed to determine the levels of tryptophan (TRP), kynurenine (KYN), kynurenic acid (KYA), 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) in rat plasma. The prepared samples were separated on a BDS column (4.6 mm × 250 mm, 5 mm) with column oven temperature of 25 °C. The mobile phase consisted of 5% acetonitrile and a buffer solution, which contained 25 mmol/L sodium acetate and 0.01 mmol/L EDTA, adjusting pH to 4.5 with acetic acid, and it was pumped at a flow-rate of 1.0 mL/min. KYN and KYA were measured by a variable wavelength detector at wavelengths 360 nm and 333 nm respectively, TRP and vanillic acid (as IS) both were measured at 280 nm. Determination of 5-HT and 5-HIAA was accomplished at the electrochemical working potential of 700 mV. Total run time was 14 min. Several parameters of the developed method were validated including linearity, accuracy precision, and stability. The results showed the established method had good LOD and separation for all of the five compounds and IS in the biological matrix. The method is simple, fast, economical and accurate. The analytical method and the results could provide a reference for the clinical and scientific research of depression.

Abnormal modification of histone acetylation involved in depression-like behaviors of rats induced by chronically unpredicted stress.

Depression is a complex multifactorial mental disorder. Its etiology involves many factors such as social environments, genetics, and psychology. Recent studies have shown that epigenetic modification may be associated with depression. Histone acetylation is one of the main mechanisms of epigenetic modification and plays an important role in genetic expression. In this study, we investigated the role of histone acetylation in the depression-like behaviors of rats undergoing chronically unpredicted stress (CUS) by detecting the mRNA and protein expression of histone deacetylase 5, cAMP-response element-binding protein, and the level of histone acetylated modification of H3K14, H3K23, and H4K16 in the prefrontal cortex and hippocampus of the rats. The results showed that significantly increasing depression-like behaviors were observed with a decreasing histone acetylated modification level, especially on acytelated-H3K14, acytelated-H3K23, and acytelated-H4K16, upregulating histone deacetylase 5 expression and downregulating cAMP-response element-binding protein expression in CUS rats, compared with control rats. The results indicate that the decrease in the histone acetylation modification level may be partly involved in the mechanism of depression-like behaviors of rats induced by CUS.

Abnormal levels of seven amino neurotransmitters in depressed rat brain and determination by HPLC-FLD.

The determination of amino acids with actions like neurotransmitters or modulators has been increasingly important for diagnosis in many neuropsychiatric diseases. A rapid and simple high-performance liquid chromatography with fluorescence detection method was developed for simultaneous determination of seven amino acids: aspartate (Asp), glutamate (Glu), serine (Ser), glutamine (Gln), glycine (Gly), taurine (Tau) and γ-aminobutyric` acid (GABA). Homoserine was used as an internal standard. The analysis was performed on a BDS column with methanol and 50 mm sodium acetate solution (pH 6.5) using a simple gradient elution. Several parameters of the developed method were validated including linearity, accuracy, precision, extraction recovery and stability, which were within the acceptable range. The method was successfully applied to determination of real samples: hippocampus and cortex in depressed rats exposed to chronically unpredictable stress in order to study if there existed differences in the seven amino acids levels between depressed rats and control. The results showed that Asp, Gly, Tau and GABA significantly decreased with increasing Gln in the hippocampus of depressed rats, compared with that of the control group, among which obviously lower level of Asp and higher level of Gln in cortex were observed. The analytical method and the results could be useful for clinical diagnosis and further insight into pathophysiological mechanism of depression.

Infliximab ameliorating depression-like behavior through inhibiting the activation of the IDO-HAAO pathway mediated by tumor necrosis factor-α in a rat model.

In recent years, some studies have suggested that the activation of inflammatory system plays a role in the occurrence of depression. Tumor necrosis factor-α (TNF-α), as one of the preinflammatory cytokines, has been reported to be involved in the occurrence of various diseases including depression. Infliximab, an antagonist of TNF-α, is usually used to treat some autoimmune diseases such as Crohn's disease and can perhaps be used to treat other diseases. In this study, the antidepressant effect and a possible mechanism of infliximab were investigated by studying the depression-like behavior and expression of TNF-α, indoleamine 2, 3-dioxygenase (IDO), and 3-hydroxyl amino acid oxygenase (HAAO) from the cortex and hippocampus in rat exposed to chronic unpredicted stress. Forty male Sprague-Dawley rats were divided into a control group (CG), an infliximab-treated control group, a model group (MG), and an infliximab-treated model group (IFXM). Infliximab (5 mg/kg once week) was administered to the infliximab-treated control group and IFXM rats by an intraperitoneal injection, whereas an equivalent volume of vehicle was administered to CG and MG rats. Rat behaviors and the expression of TNF-α, IDO, and HAAO in the cortex and hippocampus were determined. It was found that a significant relief in depression-like behaviors was observed with a downregulation of TNF-α, IDO, and HAAO expression in the IFXM rats compared with MG rats. The results show the antidepressant effect of infliximab and suggest that its mechanism is partly related to inhibition of IDO-HAAO pathway activation mediated by TNF-α in rat brain.

Simultaneous determination of eleven compounds related to metabolism of bioamines in rat cortex and hippocampus by HPLC-ECD with boron-doped diamond working electrode.

A simple and rapid high-performance liquid chromatography method with electrochemical detection employing boron-doped diamond electrode (BDD) was established for simultaneous determination of eleven bioamines with their precursor amino acids and metabolites, including two precursors (tyrosine and tryptophan), three catecholamines (dopamine, norepinephrine and epinephrine) and their four metabolites (3,4-dihydroxyphenylacetic acid, 3-methoxytyramine, homovanillic acid, and 3-methoxy-4-hydroxyphenylglycol), as well as serotonin and its metabolite (5-hydroxyindoleacetic acid), in a single run of 20 min using vanillic acid as internal standard. The separation was performed on an ODS2 column (250 mm × 4.6 mm, 5.0 µm) with column oven temperature of 30 °C. Quantification was accomplished at an oxidation potential of 700 mV vs Ag/AgCl reference electrode after a range of applied voltages were tested. Several parameters of this new chromatographic method were validated after optimizaton of the analytical conditions. The new method was successfully applied to test cortex and hippocampus samples from Sprague-Dawley rats with good separation. These eleven compounds in cortices and hippocampi were compared, which was used for monitoring their variations in neuroscience research.

Enhancing tyrosine hydroxylase and tryptophan hydroxylase expression and improving oxidative stress involved in the antidepressant effect of sodium valproate on rats undergoing chronic unpredicted stress.

Depression is a common worldwide mental disorder whose etiology remains unclear; there is also a lack of effective therapeutic agents. Sodium valproate (VPA) is a traditional antiepileptic drug with mood-stabilization effect and is increasingly being used to treat bipolar disorders and depression, but its antidepressant mechanism remains unknown. The aim of the present study was to investigate the possible mechanisms of antidepressant action by studying malondialdehyde level, catalase, and superoxide dismutase activities in the serum and the mRNA and protein expression of tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) in the prefrontal cortex of rats exposed to chronic unpredicted stress (CUS). Male Sprague-Dawley rats were used to establish a depression model by CUS. VPA (300 mg/kg once daily) and an equivalent volume of vehicle were administered to rats by an intragastric gavage. Rat behaviors, serum malondialdehyde level, serum catalase and superoxide dismutase activities, and the mRNA and protein expressions of TH and TPH in the prefrontal cortex were determined. The results showed that VPA treatment led to a significant decrease in depression-like behaviors, improvement in oxidative stress imbalance, and enhancement of TH, TPH mRNA, and protein expression in stressed rats, but failed to show any significant changes in control rats. This could indicate that the antidepressant mechanism of VPA is perhaps linked to upregulation of TH and TPH expression and inhibition of oxidative damage in CUS rats.

The Influence of Plasma Albumin Concentration on the Analysis Methodology of Free Valproic Acid by Ultrafiltration and Its Application to Therapeutic Drug Monitoring.

BACKGROUND: Free drug analysis is increasingly becoming popular in therapeutic drug monitoring (TDM). Centrifugal ultrafiltration (CF-UF) is the primary method to separate free drug from that of bound drug. However, the volume ratio of ultrafiltrate to sample solution (Vu/Vs) affects the accuracy of CF-UF, which highly depends on the different plasma conditions. Plasma protein concentrations in patients are different from those observed in healthy subjects, and there are also significant differences among patients with different diseases. Only very few studies have reported on the effect of protein concentration on the analysis methodology of free drug by CF-UF. METHODS: In this study, valproic acid was used as the representative drug, and plasma samples with different albumin concentrations were analyzed by CF-UF and hollow fiber centrifugal ultrafiltration (HFCF-UF). RESULTS: There was no significant difference of free drug concentrations by HFCF-UF and CF-UF when plasma albumin concentrations ranged 40-60 g/L. However, at low albumin concentrations (<40 g/L), a considerable difference was detected, and the difference was increased with the decrease of plasma albumin concentration. When the albumin concentration was as low as 10 g/L, the free drug concentration was 17.3 mcg/mL by CF-UF, whereas it was 10.2 mcg/mL by HFCF-UF. CONCLUSIONS: The accuracy of free drug measurement by CF-UF was albumin concentration dependent. However, such an effect was not observed when samples were prepared by HFCF-UF, which was more suitable for TDM of plasma samples from different patients. Therefore, this method could be readily applied to the measurement of free valproic acid plasma concentrations for TDM in patients.

Detection of Xeljanz enantiomers in diethyl amine active pharmaceutical ingredients and tablets.

A high-performance liquid chromatography (HPLC) method was established to detect Xeljanz enantiomers in active pharmaceutical ingredients (APIs) and tablets. The separation was achieved on a Chiralpak IC column using a mobile phase of hexane-ethanol-diethylamine (65:35:0.1, v/v). The detection wavelength was 289 nm. The peak areas and the enantiomer concentrations in the range of 0.15-2.25 µg•mL(-1) were in high linearity, with correlation coefficients higher than 0.999. The recoveries were 86.44% at the concentrations of 7.5, 18.75, and 37.5 µg•mL(-1) . The limit of detection (LOD) and limit of quantification (LOQ) were 0.042 and 0.14 µg•mL(-1) , respectively. This HPLC method is suitable for detecting the enantiomers of Xeljanz in its APIs and tablets.

Upregulating serotonin transporter expression and downregulating monoamine oxidase-A and indoleamine 2, 3-dioxygenase expression involved in the antidepressant effect of sodium valproate in a rat model.

Sodium valproate (VPA) is widely used as an antiepileptic agent and mood stabilizer. In recent years, VPA has been increasingly used as a psychotherapeutic drug to treat depression. In this article, a possible antidepressant mechanism of VPA was investigated by studying the expression and therefore the involvement of tryptophan hydroxylase, serotonin transporter (5-HTT), monoamine oxidase-A (MAO-A), and indoleamine 2, 3-dioxygenase (IDO) in rats exposed to chronic unpredicted stress. Male Sprague-Dawley rats were divided into four groups: the vehicle-treated control group (CG), the VPA-treated control group (VPAC), the vehicle-treated model group (MG), and the VPA-treated model group (VPAM). VPA (300 mg/kg once daily) was administered to VPAC and VPAM rats by means of intragastric gavage while an equivalent volume of vehicle was given to vehicle-treated CG and MG rats. Rat behavior and expression of tryptophan hydroxylase, 5-HTT, MAO-A, and IDO in the hippocampus were determined. A significant reduction in depression-like behaviors was observed with an upregulation of 5-HTT expression and a downregulation of MAO-A and IDO expression in VPAM rats, compared with MG rats. The results may suggest that the antidepressant mechanism of VPA is partly related to elevated serotonin level and its reuse in the vesicles of presynaptic nerve endings.

Histone acetylation and expression of mono-aminergic transmitters synthetases involved in CUS-induced depressive rats.

Histone acetylation has been linked to depression, the etiology of which involves many factors such as genetics, environments, and epigenetics. The aim of the present study was to investigate whether it was associated with epigenetic histone modification and gene expression of enzymes responsible for the biosynthesis of norepinephrine and serotonin in rat depression model induced by chronic unpredictable stress (CUS). Eight-week-old male Sprague-Dawley rats were exposed to CUS over 28 days. It was shown that the CUS-induced rats displayed remarked anxiety- and depression-like behavior with weakened locomotor activity in open field test and prolonged immobility in forced swimming test. Western blot revealed that CUS led to significant decrease in acetylation of H3 at Lysine 9 (K9) and H4 at Lysine 12 (K12) with obviously increasing histone deacetylases 5 (HDAC5) expression in hippocampus of CUS-induced rats. Meanwhile, there was an obviously decreased expression of tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) both at protein and mRNA levels. Administration of sodium valproate (VPA), a histone deacetylase 5 (HDAC5) inhibitor, not only significantly relieved the anxiety- and depression-like behaviors of CUS-induced rats but also clearly blunted decrease of H3(K9) and H4(K12) acetylation and expression of TH and TPH, and prevented increase of HDAC5 expression. The results indicate that there exists possible interrelation between TH and TPH gene expression and epigenetic histone acetylation in CUS-induced depressive rats, which at least partly contributes to the etiology of depression.