Responses of CH4, N2O, and NH3 emissions to different slurry pH values of 5.5-10.0: Characteristics and mechanisms.

Animal slurry storage is a significant source of greenhouse gas (GHG) and ammonia (NH3) emissions. pH is a basic but key factor that could pose great influence on gas emissions, but the simultaneous evaluation of its influence on GHG and NH3 emissions and the understanding of its underlying mechanism are not enough. In this work, pH was adjusted between 5.5 and 10.0 by a step of 0.5 unit by adding lactic acid and sodium hydroxide (NaOH) properly and frequently to the stored slurry during a 43-day storage period. The cumulative NH3 emissions were linearly correlated with the slurry pH, with R2 being 0.982. Maintaining the slurry pH at 5.5-6.0 could reduce NH3 emissions by 69.4%-85.1% compared with the non-treated group (CK). The pH ranges for maximum methane (CH4) and nitrous oxide (N2O) emissions were 7.5-8.5 and 6.5-8.5, respectively, and the slurry under pH 7.5-8.5 showed the highest GHG emissions. Acidification to pH 5.5 helped reduce the CH4, N2O, and total GHG emissions by 98.0%, 29.3%, and 81.7%, respectively; while alkalinization to pH 10.0 helped achieve the mitigation effects of 74.1%, 24.9%, and 30.6%, respectively. The Pearson's correlation factor between CH4 and the gene copy of mcrA under different pH values was 0.744 (p < 0.05). Meanwhile, the correlation factors between N2O and the gene copies of amoA, narG, and nirS were 0.644 (p < 0.05), 0.719 (p < 0.05), and 0.576 (p = 0.081), respectively. The gene copies of mcrA, amoA, narG, and nirS were maintained at the lowest level under pH 5.5. These results recommended keeping slurry pH lower than 5.5 with lactic acid can help control GHG and NH3 emissions simultaneously and effectively.

Heme peroxidases are responsible for the dehydrogenation and oxidation metabolism of harmaline into harmine.

Harmaline and harmine are ß-carboline alkaloids with effective pharmacological effects. Harmaline can be transformed into harmine after oral administration. However, enzymes involved in the metabolic pathway remain unclear. In this study, harmaline was incubated with rat liver microsomes (RLM), rat brain microsomes (RBM), blood, plasma, broken blood cells, and heme peroxidases including horseradish peroxidase (HRP), lactoperoxidase (LPO), and myeloperoxidase (MPO). The production of harmine was determined by a validated UPLC-ESI-MS/MS method. Results showed that heme peroxidases catalyzed the oxidative dehydrogenation of harmaline. All the reactions were in accordance with the Hill equation. The reaction was inhibited by ascorbic acid and excess H2O2. The transformation of harmaline to harmine was confirmed after incubation with blood, plasma, and broken blood cells, rather than RLM and RBM. Harmaline was incubated with blood, plasma, and broken cells liquid for 3 h, and the formation of harmine became stable. Results indicated an integrated metabolic pathway of harmaline, which will lay foundation for the oxidation reaction of dihydro-ß-carboline. Moreover, the metabolic stability of harmaline in blood should not be ignored when the pharmacokinetics study of harmaline is carried out.

Potential mechanisms of tremor tolerance induced in rats by the repeated administration of total alkaloid extracts from the seeds of Peganum harmala Linn.

ETHNOPHARMACOLOGICAL RELEVANCE: The seeds of Peganum harmala Linn have been widely used for the treatment of nervous, cardiovascular, gastrointestinal, respiratory, and endocrine diseases and many other human ailments. However, tremor toxicity occurs after overdose and is tolerated following multiple dosing. Thus far, little is known about the underlying mechanisms of tremors and tremor tolerance. AIM OF THE STUDY: To investigate the potential mechanisms of tremors and tremor tolerance induced in rats by the repeated administration of total alkaloid extracts from the seeds of P. harmala (TAEP). MATERIALS AND METHODS: A tremor model was induced in male Wistar rats by administering TAEP at a dose of 150 mg/kg/day. To evaluate tremor action, behavioral assessment was conducted by using a custom-built tremor acquisition and analysis system. To investigate the relationships between tremors and neurotransmitter levels in the brain, various neurotransmitters were simultaneously quantified by an ultra-performance liquid chromatography combined with electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) system, and the association between these two parameters was analyzed using Pearson correlation coefficients. To further elucidate the potential mechanisms of the alterations of neurotransmitter levels in cortical tissues, the protein expression levels of several important enzymes and transporters that are closely related to neurotransmitter levels were investigated. In addition, neuropathological analysis was conducted to assess the effect of TAEP on neurons in the brain. To further clarify the potential mechanisms of TAEP-induced neurodegeneration in the brain, c-fos was subjected to immunohistochemical analysis, and oxidative stress markers were examined. RESULTS: Tremors initially occurred in rats after the oral administration of TAEP at a dose of 150 mg/kg/day. However, they were tolerated following repeated dosing. The levels of 5-hydroxytryptamine (5-HT) and glycine (Gly) in cortical tissues were most likely associated with the tremor response. Tremor tolerance also likely resulted from the degeneration of cerebellar Purkinje cells. Furthermore, the alteration of 5-HT levels was mainly attributed to the downregulated expression of monoamine oxidase A (MAO-A). The degeneration of Purkinje neurons might have resulted from the overexpression of c-fos and increased oxidative stress in the cerebellum after the multiple dosing of TAEP. CONCLUSION: The tremor response induced by TAEP at high doses is closely related to the concentrations of 5-HT and Gly in cortical tissues. Tremor tolerance may also be attributed to the degeneration of cerebellar Purkinje cells after the repeated dosing of TAEP. Further studies should be conducted to elucidate the interaction of the alkaloids on the neurotransmitter receptors, the expression of related neurotransmitter receptors, the specific signaling pathway involved in regulating MAO-A, and the mechanism of the loss and functional recovery of cerebellar Purkinje neurons.

Improvement of Oxazolone-Induced Ulcerative Colitis in Rats Using Andrographolide.

Ulcerative colitis (UC) is usually accompanied with symptoms of abdominal pain, diarrhea, and bloody stool, which impair the quality of life of patients. Previous studies have shown that Andrographis paniculata extracts, which have andrographolide (AND) as their main compound, can relieve UC symptoms in patients. The aim of the study was to investigate the alleviating effect of AND on UC using the oxazolone (OXZ)-induced UC rat model. A total of 66 healthy male Sprague Dawley rats were used to evaluate the efficacy and mechanism of AND on UC (n = 11 per group) and grouped into control, model, SASP (sulfasalazine, positive control group, 500 mg/kg), AND-L (40 mg/kg), AND-M (80 mg/kg), and AND-H (120 mg/kg). The colonic disease activity index (DAI), colon length, spleen coefficient, pathological damage, and inflammation-related cytokine and protein expression levels were used as indices for evaluation. Results showed that the AND groups had reduced DAI and mortality, and significantly improved colon length and spleen coefficient compared with the model group. Furthermore, OXZ-induced histological injury was relieved significantly after AND treatment due to an improved crypt structure and reduced infiltration of inflammatory cells. Moreover, AND inhibited myeloperoxidase (MPO) activity and the secretion of interleukin-4 (IL-4), IL-13, and tumor necrosis factor α (TNF-α). The results of the anti-inflammatory mechanism revealed that AND blocked the signal transduction by reducing IL-4/IL-13 specific binding to IL-4 receptor (IL-4R) and inhibiting the phosphorylation of the signal transducer and activator of transcription 6 (p-STAT6). In conclusion, aside from natural plants, AND may be a candidate ingredient for UC therapy.

Ameliorative effect of deoxyvasicine on scopolamine-induced cognitive dysfunction by restoration of cholinergic function in mice.

BACKGROUND: Aerial parts of Peganum harmala Linn is used as a traditional medical herb for treatment of amnesia in Uighur medicine in China. Deoxyvasicine (DVAS) is one of the chief active ingredients in P. harmala, it possesses strong acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities in vitro, but the therapeutic effect and mechanisms on amnesia in vivo are unclear. PURPOSE: The objective of this study was to investigate the improvement effect of DVAS from P. harmala in learning and memory deficits of scopolamine-induced mice and elucidate the underlying mechanisms involved. METHODS: Mice were pretreated with DVAS (5, 15 and 45 mg/kg) and huperzine-A (0.2 mg/kg) by gavage for 7 days, and subsequently were daily intraperitoneally injected with scopolamine (1 mg/kg) to induce learning and memory deficits and behavioral performance was assessed by Morris water maze. To further evaluate the potential mechanisms of DVAS in improving learning and memory capabilities, pathological change, levels of various biochemical markers and protein expressions related to cholinergic system, oxidative stress, and neuroinflammation were examined. RESULTS: The results showed that DVAS could alleviate learning and memory deficits in scopolamine-treated mice. DVAS could regulate cholinergic function by inhibiting AChE and activating choline acetyltransferase (ChAT) activities and protein expressions. DVAS could induce brain-derived neurotrophic factor and protect hippocampal pyramidal cells against neuronal damage. DVAS also enhanced antioxidant defense via increasing the antioxidant enzyme level and activity of glutathione peroxidase, and anti-inflammatory function through suppressing tumor necrosis factor-α. Additionally, DVAS could regulate the neurotransmitters by elevating acetylcholine, 5-hydroxytryptamine, γ-aminobutyric acid and reducing 5-hydroxyindole-3-acetic acid and glutamic acid. CONCLUSION: Results illustrated that DVAS may be a promising candidate compound against amnesia via restoration of cholinergic function, regulating neurotransmitters, attenuating neuroinflammation and oxidative stress.

Potential Pharmacokinetic Drug⁻Drug Interaction Between Harmine, a Cholinesterase Inhibitor, and Memantine, a Non-Competitive N-Methyl-d-Aspartate Receptor Antagonist.

Harmine (HAR) is a beta-carboline alkaloid widely distributed in nature. It exhibits psychopharmacological effects of improving learning and memory. However, excessive dose of HAR can cause central tremor toxicity, which may be related to the glutamate system. Memantine (MEM) is a non-competitive N-methyl-d-aspartate receptor antagonist. It can be used for the treatment of Alzheimer's disease and also can block the neurotoxicity caused by glutamate. Therefore, combination of HAR and MEM would be meaningful and the pharmacokinetics investigation of HAR and MEM in combination is necessary. A ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was established and validated for the simultaneous quantitative determination of MEM, HAR and harmol (HOL), a main metabolite of HAR, in rat plasma after oral administration of HAR and MEM in combination (5.0 mg/kg of MEM combined with 20.0, 40.0, 80.0 mg/kg of HAR). The contents of HAR and HOL were determined after oral administration of HAR (20.0, 40.0 and 80.0 mg/kg), and the content of MEM was determined after oral administration of MEM (5.0 mg/kg). Blood samples were collected from each rat at 0 (pre-dose), 0.08, 0.17, 0.25, 0.33, 0.50, 0.75, 1.0, 2.0, 4.0, 8.0, 12.0 and 24.0 h after administration. The maximum peak concentration (Cmax) of MEM was obviously decreased, and the area under the plasma concentration versus time curve from zero to time t (AUC(0-t)) and mean residence time (MRT) were significantly increased after combination with HAR. The Cmax and AUC(0-t) of HAR and its metabolite HOL were increased after combination with MEM. These findings suggested that co-administration of HAR and MEM could extend their residence time in rats, and then might increase the efficacy for treatment of Alzheimer's disease. Therefore, this study will provide a basis for the rational combined application of HAR and MEM.

Subchronic toxicity and concomitant toxicokinetics of long-term oral administration of total alkaloid extracts from seeds of Peganum harmala Linn: A 28-day study in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: The seeds of Peganum harmala Linn, in which the most abundant active compounds are harmaline and harmine, have been widely used as a traditional medicine in various countries to treat a broad spectrum of diseases including asthma, cough, depression, Parkinson's and Alzheimer's diseases. However, few studies on long-term or subchronic toxicity of seeds of P. harmala were reported after overdose. AIM OF THE STUDY: To investigate the subchronic toxicity and concomitant toxicokinetics of total alkaloid extracts from seeds of P. harmala (TAEP) after oral administration for four weeks in rats. MATERIALS AND METHODS: The subchronic toxicity and concomitant toxicokinetics of TAEP were evaluated after 28-day oral administration in rats at daily dose levels of 15, 45, and 150 mg/kg. The signs of toxicity and mortality were monitored and recorded daily. The body weight and average food consumption were measured weekly. The analyses of hematology, biochemistry, urine, relative organ weights and histopathology were conducted at the termination of treatment and recovery phase. For concomitant toxicokinetics study, the plasma toxicokinetic parameters, tissue distribution, and excretion of predominant ingredients harmaline and harmine in TAEP and metabolites harmalol and harmol were tested. RESULTS: Following initial repeated exposure to high-dose (150 mg/kg/day) of TAEP excitotoxic reaction, such as tremor, was observed, but tolerated on the fourth day after multiple dosing. The significant alterations in blood glucose and lipid metabolism in liver were observed, but recovered after four weeks of drug withdrawal. The no-observed-adverse-effect level (NOAEL) of TAEP was considered to be 45 mg/kg/day under the present study conditions. There were no significant gender differences in most indexes of subchronic toxicity throughout the experimental period with the exception of food consumption and body weight. In concomitant toxicokinetics study, the alterations of dynamic characteristic for harmaline, harmine and metabolite harmol after multiple oral administration at three doses had been observed. Harmaline, harmine and metabolites harmalol and harmol were widely distributed in organs and there was no accumulation in the tissues examined. The reduction of harmaline and metabolite harmalol in brain after multiple dosing at dose of 150 mg/kg might be closely related to the tremor tolerance. The main excretory pathway for metabolites harmalol and harmol was urinary excretion via kidney. CONCLUSIONS: The results revealed that TAEP at doses of 15 and 45 mg/kg/day in rats might be safe. Excitotoxic reaction such as tremor occurred initially at dose of 150 mg/kg/day, however, the toxicity was tolerant and reversible. In addition, harmaline and harmine in TAEP had a quick absorption into blood and metabolized to harmalol and harmol, and there was no drug accumulation in the detected tissues. Further studies should be investigated to clarify the mechanisms of tremor tolerance and neurotoxicity of TAEP.

In vivo and in vitro metabolism and pharmacokinetics of cholinesterase inhibitor deoxyvasicine from aerial parts of Peganum harmala Linn in rats via UPLC-ESI-QTOF-MS and UPLC-ESI-MS/MS.

ETHNOPHARMACOLOGICAL RELEVANCE: Aerial parts of Peganum harmala Linn are a Uighur traditional medicinal herb in China used to treat amnesia, bronchial asthma, and cough. Deoxyvasicine (DVAS), a potent cholinesterase inhibitor exhibiting anti-senile dementia activity, is one of the chief active ingredients in aerial parts of P. harmala and plays a key role in mediating the pharmacological effects of P. harmala. However, the metabolic profiling and in vivo pharmacokinetic characteristics of DVAS still remain unknown. AIM OF THE STUDY: The aim of this present study was to investigate the metabolism and pharmacokinetic properties of DVAS in rats by using ultra-performance liquid chromatography combined with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UPLC-ESI-QTOF-MS) and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-ESI-MS/MS) method. MATERIALS AND METHODS: The metabolic profiling of DVAS was evaluated in vitro and in vivo by rat liver microsomes (RLMs) incubation and by rat bio-specimens, such as urine, feces, plasma, and bile, after the oral administration of 45 mg/kg DVAS. An efficient and sensitive UPLC-ESI-MS/MS method was developed and validated to simultaneously determine DVAS and its major four metabolites, namely, vasicine, deoxyvasicinone, vasicinone, and 1,2,3,9-tetrahydropyrrolo[2,1-b]quinazolin-3-ß-D-glucuronide in rat plasma. For pharmacokinetic studies, 32 Sprague-Dawley rats were randomly divided into four groups, namely, intravenous dosage group (2 mg/kg DVAS) and three oral dosage groups (5, 15, and 45 mg/kg DVAS). In addition, the activity of the components in plasma after intravenous administration of DVAS was evaluated by in vitro anti-butyrylcholinesterase (BChE) assays. RESULTS: A total of 23 metabolites were found in RLMs, plasma, urine, feces, and bile by UPLC-ESI-QTOF-MS. The metabolic pathway of DVAS in vivo and in vitro mainly involved hydroxylation, dehydrogenation, acetylation, methylation, glucuronidation, and O-sulphate conjugation, and the C-3 and C-9 sites were the main metabolic soft spots. All 23 metabolites were detected in the urine sample, and 13, 8, 22, and 6 metabolites were identified from rat feces, plasma, bile, and RLMs, respectively. The standard curves of DVAS and four metabolites in rat plasma showed good linearity in the concentration range of 0.82-524.00 ng/mL with acceptable selectivity, precision, accuracy, recovery, and stability. DVAS exhibited linear dose-proportional pharmacokinetics at doses of 5, 15, and 45 mg/kg after oral administration, and the average oral absolute bioavailability of DVAS was 47.46%. The in vitro anti-BChE assays implied that the inhibitive activities were mainly due to the different concentrations of prototype DVAS. CONCLUSIONS: DVAS can be rapidly absorbed and excreted by blood, and it is also extensively metabolized in vivo, and the anti-BChE activity in blood is mainly attributed to DVAS. These findings can lay a foundation for new drug development for DVAS.

[Effect of investment composition ratio for pure titanium crown and bridge on some mechanical properties of mould].

OBJECTIVE: To determine the optimal composition of a self-developing investment material by measuring physical and mechanical properties of mould. METHODS: L(9) (3(4)) orthogonal design was adopted. One hundred and fifty specimens with the size of 80 mm × 20 mm × 20 mm were prepared to measure the atmospheric temperature bending strength, high temperature bending strength and residual bending strength. Nine specimens with the size of 5 mm diameter 25 mm heigh were prepared to survey the thermal expansion curve from ambient temperature to 1150°C. RESULTS: Strengths were greatly affected by fine powder proportion in refractory and water/powder ratio. When the content of fine powder was 35% and water/powder ratio was 1:7.5, adequate atmospheric temperature strength and high temperature strength could be achieved. Moreover, the residual strength was moderate. The thermal extension curves of specimens in experiment group were almost similar. And the average linear expansion coefficient was (4 ∼ 5) × 10(-6)/°C. CONCLUSIONS: The three kinds of bending strength of self-developing investment material are compared with commercialized investment material for titanium casting when water/powder ratio and the content of fine powder are carefully controlled.

[Effect of investment for titanium crown and bridge on reactive layer of castings].

OBJECTIVE: To investigate the effect of self-developing investment (FUS-invest) on the reactive layer of titanium castings. METHODS: Three 10 mm x 10 mm x 1 mm pure titanium castings were founded using FUS-invest. Metallographical structure of reactive layer was observed. X-ray diffraction (XRD) and scanning electron microscope with energy-dispersive spectrum (EDS) were used to analyze the status of composition of the casting surface. Micro-Vickers hardness was measured. RESULTS: The metallograph indicated that the reactive layer was composed of coarse flake-shaped alpha phase of grains. The value of micro-Vickers hardness with the range 243 to 314 MPa had not significantly difference under the depth of 105 microm beneath the surface. The XRD pattern clearly showed the peaks of ZrO2, SiO2, Mg2TiO4 and TiO2. EDS analysis demonstrated that the main elements were Al, Si, Zr and Cl. CONCLUSION: FUS-invest is suited for pure titanium casting because of the thin reactive layer and less change to metallographical structure.

[Effects of the improved FUS-invest investment for zirconium-oxide titanium crown and bridge on the reaction layer].

OBJECTIVE: To investigate the effects of the improved FUS-invest investment for zirconium-oxide titanium crown and bridge on the reaction layer. METHODS: 10 mm x 10 mm x 1 mm titanium castings were invested. Spectrum analysis was done to its reaction layer, the metallographic examination was observed and Vickers hardness was measured. RESULTS: The reaction layer was not obvious by the metallographic examination. The layer with plenty of Si was hardly detectable. The needle crystal layer diffused distribution on the surface of the titanium matrix. The depth was about 55 microm. Spectrum analysis showed that the contents of Si and Zr decreased as the depth increased. The microhardness, which obviously decreased from the surface to the matrix, tended to be equal with the hardness of the matrix after the depth reached 75 microm from the surface. CONCLUSION: The thickness of the reaction layer of the improved FUS-invest investment for zirconium-oxide titanium crown and bridge was decreased to 55 microm from 85 microm, which was the thickness before improvement.