Bioaccessibility and bioavailability adjusted dietary exposure of cadmium for local residents from a high-level environmental cadmium region.

The critical health risks caused by cadmium (Cd) via dietary exposure are commonly assessed by detecting Cd concentrations in foods. Differently, in this study, the bioaccessibility and bioavailability of Cd in major local harvests were introduced to assess the dietary exposure of local residents from a high-level environmental Cd region. The results indicated that certain Cd was released into the digestive juice after in vitro digestion with a bioaccessibility of 20-63% for rice and 3-32% for leafy vegetables, and the released portion was partially absorbed by Caco-2 cells with a bioavailability of 2-21% for rice and 0.2-13% for leafy vegetables. The results obtained from the toxicokinetic model revealed that the predicted urinary Cd values from the estimated daily intake (EDI) of Cd, which accounted for bioaccessibility and bioavailability, were consistent with the actual measured values, and the EDIs were considerably lower than the acceptable daily intake. This suggests that the bioaccessibility and bioavailability adjusted dietary Cd exposure should be more precise. The key issues addressed in our study implores that a potential health risk cannot be neglected in people with high consumption of rice from high-level zone.

Safety and efficacy of dehydrated ethanol soaking of the operative field in the treatment of spontaneous hepatocellular carcinoma rupture.

BACKGROUND: The aim of our study was to evaluate the clinical safety and value of ethanol surgical field infiltration (ESFI), combined with distilled water peritoneal lavage (DWPL), after hepatectomy in patients with hepatocellular carcinoma (HCC) rupture. METHODS: Rat liver tissue samples were soaked in dehydrated ethanol for different soaking times, and 18 rats were assigned to three groups that underwent different soaking methods of the hepatectomy cut surface. We retrospectively reviewed 45 patients who underwent hepatectomy for treatment of ruptured HCC. Among these, EFSI combined with DWPL was used in 21 patients (DAW group), with only DWPL used in the other 24 patients (DW group). Clinical outcomes were compared between the two groups. RESULTS: For in vitro experiments, the depth of coagulation degeneration and necrosis increased with the duration of soaking. For in vivo experiments, rats in all three groups survived until postoperative day 7 without significant postoperative complication. In patients, the rate of post-operation complication was comparable between the two groups (P = 0.398), with no between-group differences in liver function levels. The incidence of peritoneal dissemination was significantly higher for DW than DAW group (P = 0.037). Kaplan-Meier test identified dehydrated ethanol treatment as a significant factor of disease-free survival (DFS) (P = 0.036). On univariate analysis, dehydrated ethanol treatment was associated with better prognostic outcomes, although it was not retained as an independent factor of patient outcome. CONCLUSIONS: Dehydrated ethanol soaking of the cut surface of the hepatectomy could potentially lower the risk of metastasis and improve the effect of hepatectomy for ruptured HCC as well as showed potential therapeutic value for intraoperative iatrogenic rupture of HCC.

Comparative pharmacokinetics of continuous and conventional intrathecal amphotericin B in rabbits.

We previously reported a new effective therapy, continuous intrathecal amphotericin B (AMB), for the treatment of cryptococcal meningitis, which had fewer side effects and complications than conventional intrathecal AMB. In this study, the pharmacokinetics of continuous intrathecal administration and conventional intrathecal AMB were compared in rabbits, providing a pharmacokinetic basis for the use of continuous intrathecal AMB therapy. The AMB concentration in the cerebrospinal fluid (CSF), sampled via an inserted cisterna magna catheter, was determined by a liquid chromatography-tandem mass spectrometry assay. The results revealed significant pharmacokinetic differences between the two groups. In the continuous intrathecal group (0.15 mg/kg/24 h), the concentration of AMB peaked 7.01 µg/ml at 4 h and then decreased to a stable level of 1.0 to 1.34 µg/ml, with no neurological impairments, while in the conventional intrathecal group (0.015 mg/kg), the drug concentration reached a peak of 3.41 µg/ml at 1 h and then decreased progressively, with fever and neurological impairments, including convulsion and paralysis. The pharmacokinetic results indicated that the continuous intrathecal AMB is a more effective and safe therapy than the conventional intrathecal AMB, with comparatively rational pharmacokinetics and fewer neurological impairments.

Analysis of the essential oils of Coriandrum sativum Using GC-MS coupled with chemometric resolution methods.

The essential oils extracted from Coriandrum sativum L. were analyzed by GC-MS coupled with chemometric resolution methods. Through the chemometric resolution methods, peak clusters were uniquely resolved into the pure chromatographic profiles and mass spectra of each component. Qualitative analysis was performed by comparing the pure mass spectra with those in the NIST 05 mass spectral library. Quantitative analysis was performed using the total volume integration method. A total of 118 constituents were detected, of which 104 were identified, accounting for 97.27% of the total content. The results indicate that GC-MS combined with chemometric resolution methods can greatly enhance the capability of separation and the reliability of qualitative and quantitative results. The combined method is an economical and accurate approach for the rapid analysis of the complex essential oil samples in Coriandrum sativum L.

[Quantitative determination of podophyllotoxin in dermal and blood microdialysis samples of rats by liquid chromatography-tandem mass spectrometry].

OBJECTIVE: To develop and validate a sensitive method for quantitative analysis of podophyllotoxin in blood and dermal microdialysis samples of rats based on liquid chromatography-tandem mass spectrometry (UFLC-MS-MS). METHODS: The microdialysis samples were prepared by liquid-liquid extraction using ethyl acetate with etoposide as the internal standard (IS). Podophyllotoxin was separated with an Agilent ZORBAX XDB-C18 column (2.1 mmx50 mm, 3.5 microm). The mobile phase consisted of acetonitrile: 10 mmol/L ammonium acetate (40:60, V/V) at a flow rate of 0.3 ml/min and the analysis was performed at the ambient temperature. The UFLC-MS/MS system was operated in the mode of multiple reaction monitoring using the electrospray ionization technique in positive mode. RESULTS: Podophyllotoxin and etoposide responses were optimized at the transitions m/z 432.7-->397.3 and 589.5-->229.5, respectively. Calibration curves were linear over the range 2.0-1000 ng/ml. The lowest limits of quantification and detection values were 2.0 ng/ml and 0.7 ng/ml, respectively. The inter- and intra-day precision and accuracy were both less than 15%. CONCLUSION: This selective and sensitive method can be used to quantity podophyllotoxin in the blood and dermal microdialysates of rats.

[Pharmacokinetic study of polydatin in Beagle dogs].

OBJECTIVE: To establish a method for detecting plasma polydatin using high-performance liquid chromatography (HPLC) and investigate the pharmacokinetics of polydatin in Beagle dogs. METHODS: HPLC-based detection of polydatin was performed on a Hypersil-BDS C18 column with a mobile phase of methanol and water (35:65) at the flow rate of 1.0 ml/min and the detection wavelength of 290 nm. The pharmacokinetic parameters of polydatin were calculated by non-compartment model statistics. RESULTS: The standard curve was linear within the range of 0.21-21.0 microg/ml (correlation coefficient of 0.9995, n=5). The average recovery was 90% with a relative standard deviation no more than 8.0%. The limit of detection of the method was 0.1 microg/ml. The pharmacokinetic parameters of polydatin at 3 doses were obtained, with T((1/2)); of 89.95-/+19.96 min, 152.15-/+55.82 min, and 202.36-/+66.75 min, and AUC0-infinity of 300.14-/+51.33 g.min.ml(-1), 751.72-/+173.97 g.min.ml(-1) and 1521.12-/+310.02 microg.min.ml(-1), respectively. CONCLUSION: The method we established allows effective detection of polydatin, whose pharmacokinetics conforms to the two-compartment model.

[Rapid identification of 22 abused drugs and organophosphorus pesticides in blood by LC-MS/MS].

OBJECTIVE: To develop a method for rapid identification of 22 abused drugs and organophosphorus pesticides in the blood. METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) in multiple-reaction monitoring mode (MRM) was employed for detecting the drugs and pesticides in the blood. The MRM database and criteria for identification were established, and ethyl acetate was used for extraction of the drugs. After 3 rounds of extractions of the blood sample (1 mL) using 2 mL ethyl acetate, the extract was vortexed for 3 min and centrifuged at 5000 r/min. Each organic phase was combined and evaporated by gentle N2 gas. The residue was re-dissolved in 100 L mobile phase, from which 5 L was taken for LC-MS/MS detection. RESULTS: The detection of the 22 target compounds could be completed within 10 min. The limit of detection of the target compound ranged from 0.03 to 6.00 ng/ml. Satisfactory results were obtained in proficiency testing program organized by China National Accreditation Service for Conformity Assessment. CONCLUSION: The method we established is rapid, selective and sensitive for detecting the 22 abused drugs and organophosphorus pesticides.

[Study on the chemical constituents of Sarcandra glabra by HPLC-ESI-MS/MS].

OBJECTIVE: To study and compare the main chemical constituents of Sarcandra glabra and qingrexiaoyanning capsules which were extracted by acetic ether. METHODS: The sample solution were analyzed by a Zorbax C18 column with a gradient mobile phase composed of acetonitrile and 0.1% formic acid solution. Both UV and mass spectrometry detector were used simutaneously, full-scan detection mode was evaluated for the identification of all LC peaks. RESULTS: We analyzed the mass spectrum of every LC peak and identified 26 molecular mass from the ion chromatogram of Sarcandra glabra extraction and 16 molecular mass from the extractions of qingrexiaoyanning capsule. 5 compounds were identified. CONCLUSION: High performance liquid chromatography-mass/mass spectrometry has special advantages on analyzing the chemical constituents of traditional Chinese medicine.

[Detection and typing of dengue virus using polymerase chain reaction and microwell plate hybridization].

OBJECTIVE: To establish a specific, sensitive and practicable method for detection and typing of dengue virus. METHODS: Based on the genomic sequence analysis of dengue virus types 1-4, 4 pairs of primers were designed. The specific capture probes of dengue virus types 1-4 were amplified using RT-PCR, cloned and sequenced before using them for precoating the microwell plate. The samples were amplified using biotin-labeled forward primer and reverse primer, and microwell plate hybridization was carried out for detection and typing of dengue virus types 1-4. RESULTS: The absorbance of the positive samples were higher than 0.5, while the average absorbance of the negative samples was lower than 0.1, with the S/N higher than 10. CONCLUSION: The method of PCR-ELISA we established for early detection and typing of all 4 dengue viruses seretypes.

[As2O3-induced permeability transition pore opening in mitochondria depends on Ca2+].

OBJECTIVE: To explore the mechanism of arsenite-induced permeability transition pore (PTP) opening and the role of Ca(2+). in As(2)O(3)-induced PTP opening. METHOD: The mitochondria were prepared from Wistar rat liver and mitochondrial swelling was assessed spectrophotometrically at 540 nm to evaluate PTP opening. The membrane potential of the mitochondria was measured with fluorescence spectrophotometry. RESULTS: PTP opening was induced with 10 micromol/L As(2)O(3) in the presence of 10 micromol/L Ca(2+), and the absorbance at 540 nm of the mitochondria did not decrease in response to exclusive treatment with 10 micromol/L As(2)O(3), or to 10 micromol/L As(2)O(3) plus 10 micromol/L Ca(2+) treatment with 0.5 mmol/L EGTA pretreatment. Treatment with As(2)O(3) at 0, 5, 10 and 20 micromol/L in the presence of 50, 20, 10 and 5 micromol/L Ca(2+), respectively, resulted in decreased absorbance at 540 nm of the mitochondria. CONCLUSION: Ca(2+) mediates As(2)O(3)-induced PTP opening. As(2)O(3) lowers Ca(2+) threshold necessary for eliciting PTP opening and thereby regulates cell apoptosis.

[Purification of a new phospholipase A2 homologue from Agkistrodon blomhoffii siniticus and its effects on gene expression of Hep3B cells].

A new PLA2 homologue was purified from Agkistrodon blomhoffii Siniticus by applying reverse phase (HPLC) C18 column. The molecular weight of PLA2 homologue is 13900Da and its purity is 97.2%. The results of N-terminal sequence analysis were HLLQFRKMIKKMTKK. All the fragments were determined with the protein-protein BLAST software of GenBank. BLAST analysis showed that the N-terminal sequences of 15 amino acids were highly homologous with the sequences of other PLA2.

[Resveratrol induces HepG2 cell apoptosis by depolarizing mitochondrial membrane].

OBJECTIVE: To investigate the effects of resveratrol on the proliferation, apoptosis, mitochondrial membrane potential and cell morphology of human liver cancer cell line HepG2. METHODS: The changes in HepG2 cell growth and proliferation in response to resveratrol treatment were evaluated by MTT assay, and resveratrol-induced apoptosis of HepG2 cells was investigated by flow cytometry. Inverted microscope and electron microscope were employed for observing morphological changes of the treated cells. The whole-cell mitochondrial membrane potential was measured in separate experiments using two fluorimetric probes, rhodamine123 and TMRE, respectively. HepG2 cells treated with rhodamine123 were analyzed by flow cytometry and cells treated with TMRE by confocal microscope. RESULTS: MTT assay showed that low concentrations of resveratrol produced no significant effect on the growth of HepG2 cells, whereas at high concentrations, resveratrol could obviously inhibit the cell growth in a time- and dose-dependent manner. Resveratrol also induced apoptosis of HepG2 cells, and after a 24-hour treatment, resveratrol caused sharp increment of the mitochondria membrane potential. CONCLUSION: Resveratrol is capable of inhibiting the proliferation of HepG2 cells and inducing cell apoptosis by depolarizing mitochondrial membrane potential.

[Purification of a new phospholipase A2 homologue from Agkistrodon blomhoffii siniticus and its effects on gene expression profile of Hep3B cells].

OBJECTIVE: To isolate and purify a new phospholipase A2 (PLA2) homologue from Agkistrodon blomhoffii siniticus and investigate its effects on the gene expression profile of Hep3B cells. METHODS: The PLA2 homologue was isolated and purified by reverse-phase high-performance liquid chromatography (HPLC) and its purity was determined also by HPLC. The relative molecular mass of the homologue was measured by electrospray ionization mass spectrum. The gene expression profile of Hep3B cells was detected with gene chip after exposure of the cells to 139 microg/ml PLA2 homologue for 12 h. RESULTS: The purity of the PLA2 homologue was 97.2%, whose relative molecular mass was 13,900. After exposure of Hep3B cells to 139 microg/ml PLA2 homologue for 12 h, 19 genes were down-regulated and 20 up-regulated in the cells. The genes showing altered expressions in response to the exposure were mainly involved in cell cycle control and DNA damage repair, cell apoptosis and senescence, production of signal transduction molecules and transcription factors, cell adhesion, angiogenesis, and tumor invasion and metastasis. CONCLUSIONS: The PLA2 homologue induces alterations in the expression of a wide variety of genes involved in the growth and metastasis of tumor cells. The results of this study provide clues for further study of the possible mechanism for the action of PLA2 homologue on Hep3B cells.

[Mechanism of opening of mitochondrial permeability transition pore induced by arsenic trioxide].

BACKGROUND & OBJECTIVE: Permeability transition pore (PTP) is central for apoptosis by acting as a good candidate pathway for the release of cytochrome c and apoptosis-induction factors from mitochondria. Arsenite may induce apoptosis via a direct effect on PTP. To characterize the exact mechanism for arsenite to induce PTP opening, the correlations of calcium-induced calcium release from mitochondria (mCICR) to As2O3-induced PTP opening and cytochrome c release from mitochondria were studied. METHODS: Mitochondria were prepared from Wistar rat livers. The effect of As2O3 on mitochondrial PTP opening was measured with ultraviolet (UV) spectrophotometer. The changes of Ca(2+) concentration were detected with UV spectrophotometer to monitor Ca(2+) -induced Ca(2+) release from mitochondria. Cytochrome c release mediated by Ca(2+) was measured with Western blot. RESULTS: As2O3 (10 micromol/L) combined with low concentration of Ca(2+) didn't induce PTP opening and cytochrome c release from mitochondria; while As2O3 (10 micromol/L) combined with high concentration of Ca(2+) induced PTP opening and cytochrome c release. When mCICR was inhibited, the effect of As2O3 and Ca(2+) on PTP opening and cytochrome c release was completely inhibited. CONCLUSION: As2O3-induced PTP opening and cytochrome c release depend on mCICR.

[High-performance liquid chromatography-electrospray ionization/tandem mass spectrometry of isofraxidin in Sarcandra glabra].

OBJECTIVE: To establish a method for determining isofraxidin concentrations in Sarcandra glabra and Qingrexiaoyanning capsules with high-performance liquid chromatographic-mass spectrometry. METHODS: Isofraxidin was extracted from Sarcandra glabra and Qingrexiaoyanning capsules with acetic ether and chloroform, respectively, and separated by isocratic reversed-phase chromatography. The mass spectrometric system was operated in multiple reaction monitoring mode. A pair of ions: precursor ion m/z 223 with product ion m/z 162 were chosen for the quantification of the analyte. RESULTS: The retention time of isofraxidin was 6.60 min, and the calibration curve was linear over a concentration range from 484 to 9 680 ng/ml. The average recovery was 96.7% and RSD 4.49%, with detection limit of 1 ng/ml. CONCLUSION: The method is rapid, selective and sensitive for determining isofraxidin in Sarcandra glabra and Qingrexiaoyanning capsules.

[Determination of eight elements in Dongjiang river sand by flame atomic absorption spectrometry].

Eight kinds of microelements (Mn, Cu, Zn, Fe, Na, Co, Mg and K) in Dongjiang river sand were detected and analysed by flame atomic absorption spectrometry(FAAS). The method is simple and rapid with good precision and accuracy, and eliminates the interference effectively. The regression equation and the correlation coefficient were calculated. Correlation coefficients were 0.994-1.000. The samples were digested with HNO3-HF-H2O2. The results showed that Dongjiang river sand contained abundant microelements. The content order is Na>K>Fe>Mg>Mn>Zn>Co>Cu with Na being the highest and Cu the lowest. This paper provides useful evidence for studying the health care effect of Dongjiang river sand on human body.

[Preparation of a composite membrane with collagen scaffold material for tissue culture].

OBJECTIVE: To prepare a composite membrane with collagen scaffold material for tissue culture. METHODS: Pure collagen membrane, collagen/hyaluronic acid (HA) membrane, collagen/chitosan membrane and collagen/chitosan/ HA membrane (abbreviated as composite collagen membrane) were prepared respectively, and their respective biodegradation time and the number of the cells proliferated on them were examined in vitro. RESULT: The biodegradation time of pure collagen, collagen/HA and composite collagen membranes in vitro was 1 380, 1 376 and 1 560 min respectively, and the number of cells proliferated on the collagen/HA membrane and the composite collagen membrane were greater than that on the pure collagen membrane. CONCLUSIONS: Addition of chitosan can decrease the biodegradation of the collagen membrane in vitro, while HA can not; HA Can contribute to cell adhesion and proliferation on the collagen membranes, but chitosan does not.

[High-performance capillary electrophoresis for determination of urinary citrate and oxalate].

OBJECTIVE: To develop a method for determining urinary citrate and oxalate using high-performance capillary electrophoresis. METHODS: Capillary electrophoresis with indirect ultraviolet (UV) detection was performed for determining urinary citrate and oxalate. Sodium hromate was used as the UV-absorbing background electrolyte. RESULTS: Citrate and oxalate had a migration time of 3.76 min and 3.14 min respectively. The calibration curves were linear within a limit of about 2 microg/ml for citrate and about 1 microg/ml for oxalate. The recovery rates of citrate and oxalate were 97.53% and 99.11% respectively. CONCLUSION: This method is simple, accurate and at low cost for determining citrate and oxalate in the urine.

[Determination of the total alkaloid content of Rhizoma coptidis in Gegenqinlian prescription].

OBJECTIVE: To determine the total content of alkaloids from Rhizoma coptidis in the traditional Chinese medicine prescription Gegenqinlian. METHODS: The extract of Gegenqinlian decoction and powder of the minipills were respectively purified with alumina column chromatography, and the total alkaloid content of Rhizoma coptidis in these preparations was determined using ultraviolet spectrophotometry at 350 nm. RESULTS: The total alkaloid content was 32.4 mg/g in the aqueous extract of Gegenqinlian decoction and 19.6 mg/g in Gegenqinlian minipills, with a recovery rate of 98.1% (RSD=2.9%). CONCLUSION: The method we employed yields accurate and stable results with much sensitivity for quality control of Gegenqinlian minipills, and also provides reference for study of Gegenqinlian prescription and other prescriptions that contain Rhizoma coptidis.

High-performance capillary electrophoresis for determining the contents of berberine, jatrorrhizine and palmatine in Gegenqinlian decoction.

OBJECTIVE: To develop a method for alkaloid isolation from the extract of Gegenqinlian decoction (a traditional Chinese herbal preparation) and content determination. METHOD: After the preparation of the Gegenqinlian decoction extract and the micropellets, the quaternary alkaloids of Coptis chinensis were isolated by high-performance capillary electrophoresis (HPCE) in a buffer solution containing 60% sodium phosphate (60 mmol/L, pH 8.0) and 40% methanol. 2-(4-hydroxyphenyl) ethylammonium chloride was used as the internal standard and the ultraviolet detection was performed at 254 nm for determining the contents of berberine, jatrorrhizine and palmatine in the extract of Gegenqinlian decoction. RESULTS: Berberine, jatrorrhizine and palmatine were successfully isolated within 6 min with recovery rates of 99.05%, 98.70% and 97.76% respectively and relative standard deviation of 2.12%, 1.85% and 1.95% respectively. CONCLUSION: HPCE is applicable for the determination and isolation of quaternary alkaloids in Coptis chinensis with high accuracy and recovery rate.