The protective activity of imperatorin in cultured neural cells exposed to hypoxia re-oxygenation injury via anti-apoptosis

Apoptosis is believed to play important roles in neuronal cell death associated with cerebral ischemia. We now provided evidence that imperatorin (IMp), the main composition of the dried root or rhizome of R Radix Angelicae Dahuricae, took advantage on oxygen glucose deprivated/reperfusion (OGD-R) SH-SY5Y cell line through neuronal apoptosis inhibition. Our data had shown that imperatorin reduced the number of apoptosis cells after OGD-R, this effect was associated with the inhibition of the apoptosis factors Bax and caspase-3, and the upregulation of anti-apoptosis factor Bcl-2. In the meantime, the protective factor BDNF was upregulated significantly by imperatorin treatment. In our experiment in vivo, imperatorin decreased the infract volume significantly in dose of 5 mg/kg and 10 mg/kg, and the behavior ability was increased in the 10 mg/kg of imperatorin. Our observations show that imperatorin exerted protective effect on cerebral ischemia both in vitro and in vivo, this effect is associated with its anti-apoptosis function.     

The inhibition of resveratrol to human skin squamous cell carcinoma A431 xenografts in nude mice

Squamous cell carcinoma (SCC) is one of the commonest dermatological malignancies. Resveratrol (Res) is one type of polyphenolic compound which was first identified from the roots of Veratrum grandinorum in 1940. The previous studies found that Res can promote apoptosis of a variety of tumor cell, especially SCC cells. However it is rare to study the inhibition mechanism of Res in the animal model. In this study, through the establishment of human cutaneous SCC A431 xenografts in nude mice, we observed Res inhibition effect and investigated the inhibition mechanism by checking the expression of apoptosis-related factors, p53, ERK and survivin. The results showed that the xenograft volume and weight of Res groups were less than those of the control groups (P < 0.05), but the net body mass of nude mice of Res groups was not significantly different from the control groups (P > 0.05). The apoptotic index of Res groups were significantly higher than the control groups (P < 0.05). The protein and mRNA expression of p53 and ERK were statistically positively correlated (P < 0.05) and significantly increased in Res high- and medium-dose groups compared with the control groups (P < 0.05). Moreover, the protein and mRNA expression of SVV were negatively correlated with p53 (P < 0.05) and lower than the control groups (P < 0.05). The results demonstrate Res inhibitory effect and indicate that the inhibition mechanism of Res is to upgrade the protein and mRNA expression of p53 and to downgrade the protein and mRNA expression of SVV, thus inducing the apoptosis of tumor cells.     

Emodin opposes chronic unpredictable mild stress induced depressive-like behavior in mice by upregulating the levels of hippocampal glucocorticoid receptor and brain-derived neurotrophic factor

Emodin, the major active component of Rhubarb, has shown neuroprotective activity. This study is attempted to investigate whether emodin possesses beneficial effects on chronic unpredictable mild stress (CUMS)-induced behavioral deficits (depression-like behaviors) and explore the possible mechanisms. ICR mice were subjected to chronic unpredictable mild stress for 42 consecutive days. Then, emodin and fluoxetine (positive control drug) were administered for 21 consecutive days at the last three weeks of CUMS procedure. The classical behavioral tests: open field test (OFT), sucrose preference test (SPT), tail suspension test (TST) and forced swimming test (FST) were applied to evaluate the antidepressant effects of emodin. Then plasma corticosterone concentration, hippocampal glucocorticoid receptor (GR) and brain-derived neurotrophic factor (BDNF) levels were tested to probe the mechanisms. Our results indicated that 6 weeks of CUMS exposure induced significant depression-like behavior, with high, plasma corticosterone concentration and low hippocampal GR and BDNF expression levels. Whereas, chronic emodin (20, 40 and 80 mg/kg) treatments reversed the behavioral deficiency induced by CUMS exposure. Treatment with emodin normalized the change of plasma corticosterone level, which demonstrated that emodin could partially restore CUMS-induced HPA axis impairments. Besides, hippocampal GR (mRNA and protein) and BDNF (mRNA) expressions were also up-regulated after emodin treatments. In conclusion, emodin remarkably improved depression-like behavior in CUMS mice and its antidepressant activity is mediated, at least in part, by the up-regulating GR and BDNF levels in hippocampus.     

Individual and combined effects of rhynchophylline and ketamine on proliferation,NMDAR1 and GluA2/3 protein expression in PC12 cells

Rhynchophylline is an active component of the Uncaria species, which is a member of the Rubiaceae family. Our studies show that the downregulation of N-methyl-d-aspartate (NMDA) receptor subunit GluN2B expression in the nucleus accumbens, amygdala, medial prefrontal cortex, and hippocampal CA1 area by rhynchophylline is beneficial for the treatment of psychological dependence on amphetamines. The individual and combined effects of rhynchophylline and ketamine on proliferation and GluN1 and GluA2/3 protein expression in PC12 cells were investigated. PC12 cells were differentiated into neuron-like cells by treatment with nerve growth factor (50 ng/mL). After treatment for 48 h, differentiated PC12 cell proliferation and GluN1 and GluA2/3 protein expression were analyzed. The viability of PC12 cells was reduced by ketamine at doses of 0.50, 1.00, 1.50, and 2.00 mmol/L, with the viability of cells treated with 1.50 and 2.00 mmol/L of ketamine significantly lower than that of the control cells. However, PC12 cells treated with rhynchophylline showed no toxicity at doses of 0.25, 0.50, 0.75, or 1.00 mmol/L. While GluA2/3 protein expression was upregulated by ketamine, it was not influenced by rhynchophylline. GluN1 protein expression was downregulated by rhynchophylline (1 mmol/L), while treatment with ketamine, either alone or with rhynchophylline, had no effect. These findings demonstrate that rhynchophylline suppresses GluA2/3 expression in ketamine-induced PC12 cells and downregulates GluN1 expression. Ketamine's lack of effect on GluN1 expression offers a partial explanation for ketamine addiction and the anti-addictive properties of rhynchophylline.     

Effects of rhynchophylline on GluN1 and GluN2B expressions in primary cultured hippocampal neurons

N-methyl-d-aspartate (NMDA) receptor subunits GluN1 and GluN2B in hippocampal neurons play key roles in anxiety. Our previous studies show that rhynchophylline, an active component of the Uncaria species, down-regulates GluN2B expression in the hippocampal CA1 area of amphetamine-induced rat. The effects of rhynchophylline on expressions of GluN1 and GluN2B in primary hippocampal neurons in neonatal rats in vitro were investigated. Neonatal hippocampal neurons were cultured with neurobasal-A medium. After incubation for 6 h or 48 h with rhynchophylline (non-competitive NMDAR antagonist) and MK-801 (non-competitive NMDAR antagonist with anxiolytic effect, as the control drug) from day 6, neuron toxicity, mRNA and protein expressions of GluN1 and GluN2B were analyzed. GluN1 is mainly distributed on neuronal axons and dendritic trunks, cytoplasm and cell membrane near axons and dendrites. GluN2B is mainly distributed on the membrane, dendrites, and axon membranes. GluN1 and GluN2B are codistributed on dendritic trunks and dendritic spines. After 48 h incubation, a lower concentration of rhynchophylline (lower than 400 μmol/L) and MK-801 (lower than 200 μmol/L) have no toxicity on neonatal hippocampal neurons. Rhynchophylline up-regulated GluN1 mRNA expression at 6 h and mRNA and protein expressions at 48 h, but down-regulated GluN2B mRNA and protein expressions at 48 h. However, GluN1 and GluN2B mRNA expressions were down-regulated at 6 h, and mRNA and protein expressions were both up-regulated by MK-801 at 48 h. These findings show that rhynchophylline reciprocally regulates GluN1 and GluN2B expressions in hippocampal neurons, indicating a potential anxiolytic property for rhynchophylline.     

Hormone-sensitive lipase is involved in the action of hydroxysafflor yellow A (HYSA) inhibiting adipogenesis of 3T3-L1cells

BackgroundSafflor yellow A (SY) has been demonstrated to be beneficial to cardiovascular system. Our previous study showed that hydroxysafflor yellow A (HSYA), a main component of SY, could increase peroxisome proliferator-activated receptor γ mRNA expression. In this study, we investigate the effect of HSYA on the proliferation and adipogenesis of mouse 3T3-L1 preadipocytes.MethodsThe proliferation and adipogenesis of 3T3-L1 cells treated with HSYA was studied by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) spectrophotometry, Oil Red O staining and intracellular triglyceride assay methods. HSL mRNA expression and promoter activity were studied by real-time quantitative RT-PCR, transient transfection and dual luciferase reporter gene methods.ResultsHSYA (0.1 mg/L) significantly inhibited the proliferation of 3T3-L1 cells when compared with control cells in 8 h. This effect was further enhanced with the extension time (24 to 96 h) and an increase of concentration of HSYA (1–10 mg/L). The maximal inhibitory action was observed at 0.1 mg/L HSYA in 72 h (86 ± 11.8% vs. 100 ± 4.1%, p < 0.01). HSYA notably reduced the amount of intracellular lipid and triglyceride content in adipocytes to 85% (1 mg/L) and 75% (100 mg/L) on Day 4 following the differentiation, respectively, while increased HSL mRNA expression and promoter activities to 2.7 fold and 1.55 fold, respectively (p < 0.01), in differentiated 3T3-L1 adipocytes.ConclusionsHSYA inhibits the proliferation and adipogenesis of 3T3-L1 preadipocytes. The inhibitory action of HYSA on adipogenesis may be due to the promotion of lipolytic-specific enzyme HSL expression by increasing HSL promoter activity.     

Thermal stability of ginkgolic acids from Ginkgo biloba and the effects of ginkgol C17:1 on the apoptosis and migration of SMMC7721 cells

Ginkgolic acids are alkylsalicylic acid derivatives with a thermolabile carboxylic group from Ginkgo biloba L., and they exhibit anticancer activity. Their anticancer effects are closely associated with their thermal stability. In this study, the thermal decomposition of ginkgolic acids was analyzed at temperatures of 30, 50, 70 and 250 °C. The results clearly showed that an obvious slow decarboxylation of the ginkgolic acids was detected at 70 °C. When the temperature increased to 250 °C, the decarboxylation reaction was rapidly completed. The ginkgolic acids were decarboxylated to yield ginkgols. The ginkgols C13:0, C15:1 and C17:1 were separated and definitively identified by IR, NMR and GC-MS. The cytotoxic effects of ginkgols C13:0, C15:1 and C17:1 were tested and compared with those of the corresponding ginkgolic acids. An MTT assay showed that ginkgol C17:1 (48-h IC₅₀ = 8.5 μg · ml^- 1) has the strongest inhibition on SMMC-7721 cells in a dose- and time-dependent manner. The anticancer action may occur via the induction of apoptosis by the activation of caspases-3, the upregulation of Bax expression, and the inhibition migration of SMMC7721 cells. The results indicated that ginkgol C17:1 might be useful for the further development of a hepatocellular carcinoma preventive agent.     

Fine-root distribution and morphology in an acidic Norway spruce (Picea abies (L.) Karst.) stand in SW Sweden in relation to granulated wood ash application

Wood ash is recommended as a compensatory fertiliser to counteract the effects of acidic deposition on forest ecosystems. Spatial distribution of biomass, necromass and morphology parameters of the fine roots (diameter classes <1, 1–2, <2 mm) of Norway spruce (Picea abies (L.) Karst.) were analysed in response to fertilisation with granulated wood ash (GWA) in a long-term field experiment in SW Sweden. GWA was applied as a single dose of 3200 kg ha⁻¹ and the fine roots were sampled 9 years later by soil coring. Soil cores were divided into 1-cm strata within the top 0–2.5 cm humus limits, the lower humus below 2.5 cm (with varying thickness) and the mineral soil to 50 cm depth (from ground surface). Total fine-root biomass in the control (C) and GWA treatment, 256 ± 20 and 258 ± 25 g m⁻², respectively, and length 2072 ± 182 and 1800 ± 198 m m⁻², respectively, did not differ statistically from each other. Total fine-root necromass in the 1–2 mm fraction was significantly higher in C than in the GWA treatment, 130 ± 12 and 80 ± 10 g m⁻², respectively. Fine-root biomass in the <1 mm fraction was significantly greater in the lower humus in the GWA treatment, but this did not affect the total biomass in the <1 mm fraction in the whole soil profile. The biomass-to-necromass ratio (1–2 mm) was significantly higher in the GWA treatment in the 0–30 cm soil layer than in the corresponding layer of the control. Specific root length (SRL) was lower in the GWA treatment than in the control in the 0–5 cm soil layer. The lower necromass and SRL were more clearly related to the GWA treatment, whereas the difference in the vertical distribution of biomass may have been related to the thicker humus layer in the GWA plots.     

Baicalein decreases side population proportion via inhibition of ABCG2 in multiple myeloma cell line RPMI 8226 in vitro

ObjectiveTo investigate the effect of baicalein on side population in human multiple myeloma cell line RPMI 8226 and the underlying molecular mechanisms in vitro and in silico.MethodsMTT assay was applied to detect the anti-proliferation effect of baicalein. The detection of side population cells is based on the Hoechst 33342 exclusion assay technique and flow cytometric analysis. Western blotting assay was used to explore the expression of ABCG2 protein. Homology modeling and molecular docking were performed with Discovery Studio 2.1.ResultsBaicalein decreased both cell viability with IC₅₀ = 168.5 μM and the proportion of SP cells in a dose-dependent manner. Correspondingly, it significantly decreased the expression level of ABCG2 protein. Baicalein also shared similar binding sites and modes with fumitremorgin C to the protein.ConclusionsBaicalein possessed novel anticancer properties, such as anti-proliferation and drug efflux inhibition in side population cells, which suggested its potential feature of targeting cancer stem cells of multiple myeloma.     

Stand development after different thinnings in two uneven-aged Picea abies forests in Sweden

Two field experiments, located in Central and Northern Sweden, were used to study the influence of standing volume on volume increment and ingrowth in uneven-aged Norway spruce (Picea abies (L.) Karst.) stands subjected to different thinnings. Each experiment had a 3 × 2 factorial block design with two replications. Treatments were thinning grade, removing about 45, 65, and 85% of pre-thinning basal area, and thinning type, removing the larger or the smaller trees, respectively. Each site also had two untreated control plots. Plot size was 0.25 ha. Volume increment was 0.5–6.8 m³ ha⁻¹ year⁻¹ for the plots, and significantly positively (p < 0.01) correlated with standing volume. Within treatment pairs, plots thinned from Above had consistently higher volume increment than plots thinned from Below. Ingrowth ranged from 3 to 33 stems ha⁻¹ year⁻¹, with an average of 14 and 21 stems ha⁻¹ year⁻¹ at the northern and southern site, respectively. At the southern site ingrowth was significantly negatively (p < 0.01) correlated with standing volume, but not at the northern site. Mean annual mortality after thinning was 2 and 7 stems ha⁻¹ year⁻¹at the northern and southern site, respectively.     

Structural characterization and antioxidant activity of a low-molecular polysaccharide from Dendrobium huoshanense

The present study aimed at investigating the structural features and antioxidant activities of a polysaccharide fraction (DHP1A) obtained from Dendrobium huoshanense, a precious herb medicine in China. DHP1A mainly consisted of mannose (Man), glucose (Glc) and a trace of galactose (Gal), with a molecular weight of 6700 Da. Its backbone contained (1 → 4)-linked α-D-Glcp, (1 → 6)-linked α-D-Glcp and (1 → 4)-linked β-D-Manp, with a branch of terminal β-D-Galp. The in vitro antioxidant evaluation revealed that DHP1A had a remarkable inhibition effect on the FeCl₂-induced lipid peroxidation. Furthermore, DHP1A pretreatment decreased the production of malondialdehyde (MDA), and restored the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), as well as the level of glutathione (GSH) in the livers of CCl₄-treated mice. These results suggested that DHP1A was a potential antioxidant component in D. huoshanense.     

Utilization and traditional strategies of in situ conservation of iroko (Milicia excelsa Welw. C.C. Berg) in Benin

Socio-cultural surveys were carried out on the basis of a questionnaire administered on 346 respondents in order to investigate cultural and ethnobotanic uses of Milicia excelsa in Benin.M. excelsa contributes to cure 45 human diseases. The different parts of the tree used are leaves (30.3%), bark (25.8%), root (23.6%), latex (10.1%), flaking bark (6.7%), wood, calcium concentrated in old trees, and gum (1.1% each). Fruits or seeds are rarely used. Six different forms of utilization were recorded: soaking (46.3%), bark or leaves decoction (32.8%), herb tea (11.9%), powder (6.0%), leaves or bark grounded and rolled up into ball (1.5%), component of offering to fetish (1.5%). Iroko wood is also used in carpentry and joinery for construction purposes; furniture as well as for building boats/canoes.Iroko tree is used as the conservatory of cultural values and incarnates many divinities, which differ significantly from one province to another (χ² = 1830.27; d.f. = 25; P < 0.01%). There is a significant difference between the provinces in respect of the recognition of the species (χ² = 268.71; d.f. = 17; P ≤ 0.01%) and the population awareness about iroko as a sacred tree also varied from one province to another (χ² = 308.66; d.f. = 27; P ≤ 0.01%).Veneration of the tree is the main approach of its conservation by local people. M. excelsa is conserved on farm, in sacred groves, in public places and in cemeteries. The different sacred objects used to symbolize the divinities incarnated by iroko are: pottery (36.36%), iron (11.11%), calabash (4.04%), candle (2.02%), piece of cloth (18.18%), sacrifice (13.13%), piece of money (3.0%), stone (2.05%), glassware (broken bottle, 2.02%), and convent (8.08%). There is a highly significant difference between provinces as far as the sacred objects are concerned (χ² = 183.037; d.f. = 19 and P < 0.001%). The conservation purposes also vary significantly from one region to other (χ² = 894.47; d.f. = 31; P < 0.01%).     

Characteristics of nobiletin-induced effects on jejunal contractility

Nobiletin, a citrus polymethoxylated flavone, exhibits multiple biological properties including anti-inflammatory, anti-carcinogenic, and anti-insulin resistance effects. The present study found that nobiletin exerted significant stimulatory effects on the contractility of isolated rat jejunal segments in all 6 different low contractile states, and meanwhile significant inhibitory effects in all 6 different high contractile states, showing characteristics of bidirectional regulation (BR). Nobiletin-exerted BR on jejunal contractility was abolished in the presence of c-kit receptor tyrosine kinase inhibitor imatinib or Ca^2 + channel blocker verapamil. In the presence of neuroxin tetrodotoxin, nobiletin only exerted stimulatory effects on jejunal contractility in both low and high contractile states. Hemicholinium-3 and atropine partially blocked nobiletin-exerted stimulatory effects on jejunal contractility in low-Ca^2 +-induced low contractile state. Phentolamine or propranolol or l-NG-nitro-arginine significantly blocked nobiletin-exerted inhibitory effects on jejunal contractility in high-Ca^2 +-induced high contractile state respectively. The effects of nobiletin on myosin light chain kinase (MLCK) mRNA expression, MLCK protein content, and myosin light chain phosphorylation extent were also bidirectional. In summary, nobiletin-exerted BR depends on the contractile states of rat jejunal segments. Nobiletin-exerted BR requires the enteric nervous system, interstitial cell of Cajal, Ca^2 +, and myosin phosphorylation-related mechanisms.     

(RS)-glucoraphanin purified from Tuscan black kale and bioactivated with myrosinase enzyme protects against cerebral ischemia/reperfusion injury in rats

Ischemic stroke is the result of a transient or permanent reduction in cerebral blood flow caused by the occlusion of a cerebral artery via an embolus or local thrombosis. Restoration of blood supply to ischemic tissues can cause additional damage known as reperfusion injury that can be more damaging than the initial ischemia.This study was aimed to examine the possible neuroprotective role of (R_S)-glucoraphanin, bioactivated with myrosinase enzyme (bioactive R_S-GRA), in an experimental rat model of brain ischemia/reperfusion injury (I/R). R_S-GRA is a thiosaccharidic compound found in Brassicaceae, notably in Tuscan black kale (Brassica oleracea L. var. acephala sabellica).The mechanism underlying the inhibitory effects of bioactive R_S-GRA on inflammatory and apoptotic responses, induced by carotid artery occlusion in rats, was carefully examined. Cerebral I/R was induced by the clamping of carotid artery for 1 h, followed by 40 min of reperfusion through the release of clamp.Our results have clearly shown that administration of bioactive R_S-GRA (10 mg/kg, i.p.) 15 min after ischemia, significantly reduces proinflammatory parameters, such as inducible nitric oxide synthase expression (iNOS), intercellular adhesion molecule 1 (ICAM-1), nuclear factor (NF)-kB traslocation as well as the triggering of the apoptotic pathway (TUNEL and Caspase 3 expression).Taken together our data have shown that bioactive R_S-GRA possesses beneficial neuroprotective effects in counteracting the brain damage associated to I/R. Therefore, bioactive R_S-GRA, could be a useful treatment in the cerebral ischemic stroke.     

Initial impact of supervised logging and pre-logging climber cutting compared with conventional logging in a dipterocarp rainforest in Sabah,Malaysia

Pre-marked skid trails, directional felling and climber cutting when logging in tropical rainforests may be important ways of reducing damage to the forest, thus creating a healthier stand and improving future yields.This study, carried out in a virgin dipterocarp rainforest in the south of Sabah, Malaysia, compared two types of logging (both with and without pre-cutting climbers): conventional selective logging (CL) and supervised logging (SL). The latter is a selective logging system in which both pre-marked skid trails and directional felling were implemented. The pre-marked skid trails were aligned parallel to each other, spaced 62 m apart. A randomised complete block 2 × 2 factorial design was used in the experiment, consisting of 16 gross treatment plots, each of 5.76 ha with a 1 ha net plot in the centre.Fewer trees tended (0.050 < P ≤ 0.100) to be logged in SL plots than in CL plots (on average 9.4 and 13.0 trees ≥60 cm diameter breast height ha⁻¹). Pre-felling of climbers resulted in four more dipterocarp trees being logged ha⁻¹, compared with no climber cutting: a statistically significant difference (P ≤ 0.050). The basal areas lost of both large trees (≥ 60 cm dbh) and small dipterocarp trees (10–29 cm dbh) tended to differ between the logging systems, with CL leading to greater losses.There were significant differences in the residual stands left by the logging systems, with respect to the number of dipterocarps and their basal area in the diameter class 10–29 cm; ca 30% more stems being found after SL. No significant differences (or tendencies) in these variables were found in the residual stands in other diameter classes, or when trees of all species were considered.     

Dynamics of above- and below-ground biomass and nutrient accumulation in an age sequence of Nothofagus antarctica forest of Southern Patagonia

Nothofagus antarctica (Forster f.) Oersted is a deciduous tree species, which naturally grows on poorly drained or drier eastern sites in the Andes Mountain near Patagonian steppe. Above- and below-ground biomass and nutrients pools were measured in pure even-aged stands at different ages (5–220 years) and crown classes. Functions were fitted for total biomass and nutrients accumulation, and root/shoot ratio of individual trees against age. Total biomass accumulated for mature dominant trees was eight times greater than mature suppressed trees. Biomass root/shoot ratio decreased with age from 1.8 to a steady-state of 0.5. All nutrients concentration (except Ca) decreased with age and varied according to the degree of crown suppression classes. Nutrient concentrations varied between biomass pool components following the order leaves > bark > small branches > fine roots > medium roots > rooten wood > coarse roots > sapwood > heartwood. Total nutrient accumulation followed the order dominant > codominant > intermediate > suppressed trees and its accumulation rate varied over time, e.g. P accumulation rate of dominant trees increased from 0.17 g tree⁻¹ year⁻¹ during regeneration to 1.39 g tree⁻¹ year⁻¹ in mature trees. Nutrients uptake reached a peak during the period of maximum biomass production, and root/shoot ratio of nutrients decreased from its maximum value at 5 years of age (0.6, 4.0, 0.9, 1.5, 1.0 and 2.6 for N, P, K, Ca, S and Mg, respectively) to a steady-state asymptote beyond 50 years of age. Thus, accumulation of nutrients in roots was greater during the regeneration phase of stand development, and nutrient accumulation increased in above-ground over time. Also, nutrient use efficiency increased in mature trees (111–220 years) and decreased in suppressed crown classes. The equations developed for individual trees have been used to estimate stand biomass and nutrient accumulation from forest inventories data. Total stand biomass varied from 62.5 to 133.4 t ha⁻¹ and total nutrients accumulation ranged from 3 kg Mg ha⁻¹ to 1235 kg Ca ha⁻¹. Proposed equations can be used for practical purposes such as to estimate pasture nutrients requirement in a silvopastoral system based on nutrients supply from leaf litter returns, or to determine amelioration practices like debarking stems before harvesting.     

Pharmacokinetic and metabolism studies of rohitukine in rats by high performance liquid-chromatography with tandem mass spectrometry

A sensitive, selective, and rapid high performance liquid chromatography–tandem mass spectrometry (LC–MS/MS) was developed for the quantification of rohitukine in rat plasma. HPLC was performed using a Symmetry-Shield C₁₈ (5 μ, 4.6 × 150 mm) column, and isocratic elution with ammonium acetate buffer (pH 4; 10 mM):methanol (08:92, v/v) at a flow rate of 0.6 mL/min. Sample clean-up involved solid phase extraction (SPE) of analyte and internal standard (phenacetin) from 100 μL plasma. The parent → product ion transitions (MRM) for analyte and IS were 306.1 → 245.1 m/z and 180.1 → 138.1 m/z respectively, and were monitored on a triple quadrupole mass spectrometer, operating in positive ion mode. The method was validated across the dynamic concentration range of 5–500 ng/mL for rohitukine, with a fast run time of 4.5 min. The analytical method measured concentrations of rohitukine with accuracy (% bias) of <± 10% and precision (% RSD) of <± 12%. Rohitukine was stable during the battery of stability studies viz., bench-top, auto-sampler, freeze/thaw cycles and 30 days of storage in a freezer at − 70 ± 10 °C. Finally, the applicability of this assay has been successfully demonstrated in vivo pharmacokinetic and in vitro metabolism studies in Sprague–Dawley rat. This method will therefore be highly useful for future preclinical and clinical pharmacokinetic studies of rohitukine.     

Determination of oxymatrine and its active metabolite matrine in human plasma after administration of oxymatrine oral solution by high-performance liquid chromatography coupled with mass spectrometry

A rapid, sensitive and selective high-performance liquid chromatography mass spectrometric method has been developed and validated for the simultaneous determination of oxymatrine and its active metabolite matrine in human plasma after administration of oxymatrine oral solution. Analytes were extracted from the plasma by liquid-liquid extraction with chloroform. The chromatographic separation was accomplished on a Venusil C₁₈ column (150 mm × 4.6 mm, 5 μm) protected by a C₁₈ guard column (4.0 mm × 2.0 nm; Phenomenex, Torrance, CA, USA). Analytes were detected on a single quadruple mass spectrometer by selected ion monitoring mode via electrospray ionization source. The assay had a lower limit of quantification of 1.5 ng · mL^− 1 for oxymatrine and 3 ng·mL^− 1 for matrine in plasma. The calibration curves were linear in the measured range. The overall precision and accuracy for all concentrations of quality controls and standards were within ± 15%. The proposed method enabled unambiguous identification and quantification of oxymatrine and its active metabolite matrine in vivo. The results provided a meaningful basis for evaluating the clinical applications of the oxymatrine oral solution.     

Protection of chronic renal failure by a polysaccharide from Cordyceps sinensis

A water-soluble polysaccharide (CPS-2), isolated from the cultured Cordyceps sinensis, was obtained by hot-water extraction, anion-exchange and gel permeation chromatography. Its structural characteristics were investigated by PMP pre-column derivation, periodate oxidation, methylation analysis, FTIR and NMR spectroscopy. CPS-2 was found to be mostly of α-(1 → 4)-d-glucose and α-(1 → 3)-D-mannose, branched with α-(1 → 4,6)-d-glucose every twelve residues on average. CPS-2 had a molecular weight of 4.39 × 10⁴ Da. The protective effect of CPS-2 on the model of chronic renal failure was established by fulgerizing kidney. The changes in blood urea nitrogen and serum creatinine revealed that CPS-2 could significantly relieve renal failure caused by fulgerizing kidney.