A thiol-selective and acid-stable protein modification strategy using an electron-deficient yne reagent.

A protein modification strategy was developed based on a thiol-yne click reaction using an electron-deficient yne reagent. This approach demonstrated exceptional selectivity towards thiols and exhibited rapid kinetics, resulting in conjugates with superior acid stability. The conjugation of IgG with an indole-derived fluorophore was achieved for the imaging of PD-L1 in cancer cells.

LIMD2 promotes tumor proliferation, invasion, and epithelial-mesenchymal transition in clear cell renal cell carcinoma.

LIMD2 was found upregulated in various tumors and metastatic samples and associated with a poor prognosis. But the role of LIMD2 in clear cell renal cell carcinoma (ccRCC) remains elusive. The expression of LIMD2 in ccRCC was analyzed using cohort data downloaded from TCGA and ICGC databases. In vitro and in vivo experiments were then conducted to study the biological role of LIMD2 in ccRCC and explore the possible mechanism. The results indicated that LIMD2 was overexpressed and correlated with a poor outcome in ccRCC. LIMD2 promoted the malignancy of ccRCC both in vitro and in vivo. LIMD2 induced epithelial-mesenchymal transition (EMT) via activating the ILK/Akt pathway in ccRCC. In conclusion, LIMD2 is overexpressed and promotes proliferation, invasion, and EMT in ccRCC, which may serve as a potential novel therapeutic target for ccRCC.

[Effect of Pien Tze Huang against EV71].

This study aims to investigate the inhibitory effect of Pien Tze Huang(PZH) on enterovirus 71(EV71). To be speci-fic, chemiluminescence method was adopted to evaluate the toxicity of PZH to African green monkey kidney(Vero) cells and human rhabdomyosarcoma(RD) cells, and cytopathic effect(CPE) method to assess the inhibition on EV71-GFP reporter virus and EV71 C4 wild-type virus. The results showed that PZH had low cytotoxicity to Vero cells and RD cells, with the half-maximal cytotoxic concentration(CC_(50)) of about 0.691 3-0.879 2 mg·mL~(-1) for the two. In addition, PZH can effectively inhibit the replication of EV71 within the non-cytotoxic concentration range, and dose-dependently alleviate the cytopathic changes caused by virus infection, with the half-maximal effective concentration(EC_(50)) of 0.009 2-0.106 3 mg·mL~(-1). On the basis of the above results, the green fluorescent protein(GFP), indirect immunofluorescence assay(IFA), and median tissue culture infective dose(TCID_(50)) were employed to assess and verify the anti-EV71-GFP and anti-EV71 C4 activity of PZH. The results demonstrated that PZH can dose-dependently lower the expression of GFP by EV71-GFP and structural protein VP-1 by EV71 C4 and decrease the production of progeny infectious viruses. The EC_(50) of PZH for EV71-GFP and EV71 C4 was about 0.006 0-0.006 2 mg·mL~(-1) and 0.006 6-0.025 6 mg·mL~(-1), respectively. This study suggested that PZH may exert antiviral activity by acting on EV71 and interfering with the expression of VP-1. At the moment, there is still a lack of specific anti-EV71 drugs. This study proposed a new idea for the symptomatic treatment of EV71 infections such as hand-foot-mouth disease and verified an effective drug for the treatment of EV71 infections.

Complete duodenal obstruction induced by groove pancreatitis: A case report.

BACKGROUND: Groove pancreatitis (GP) is a type of chronic pancreatitis occurring in an anatomic area between the duodenum, head of the pancreas, and common bile duct. Duodenal obstruction is always caused by malignant pancreatic diseases, such as pancreatic head carcinoma, while is rarely induced by benign pancreatic diseases, such as pancreatitis. CASE SUMMARY: A 39-year-old man presented with a 1-mo history of upper abdominal discomfort. His concomitant symptoms were abdominal distension, postprandial nausea, and vomiting. Contrast-enhanced computed tomography of the abdomen showed thickening of the intestinal wall with enhancement of the descending segment of the duodenum, which could not be clearly differentiated from the head of the pancreas. Upper gastrointestinal radiographs and gastrointestinal endoscopy showed a complete obstruction of the descending duodenum. An operation found that a 3-cm mass was located in the "groove part" of the pancreas and oppressing the descending duodenum. Pancreaticoduodenectomy was performed to relieve the obstruction and thoroughly remove the pancreatic lesions. The pathologic diagnosis was pancreatitis. The patient had an uneventful recovery with no complications. CONCLUSION: Because of the special location and the contracture induced by long-term chronic inflammation, our case reminds surgeons that some benign pancreatic diseases, such as GP, can also present with symptoms similar to those of pancreatic cancer. This knowledge can help to avoid an unnecessary radical operation.

[Effect of Iron on the Release of Arsenic in Flooded Paddy Soils].

Amorphous iron oxides in paddy soil are critical adsorbents of arsenic. The flooding period during rice cultivation contributes to the reductive dissolution of these amorphous iron oxides, which releases sorbed arsenic into the paddy soil solution. However, more detailed work should be conducted to evaluate quantitatively arsenic immobilization, release, and transformation regulated by metastable amorphous iron oxides. In previous studies, arsenic in the soil solution phase and solid phase were classified into F1 (exchangeable arsenic), F2 (specifically sorbed arsenic), F3 (amorphous iron oxide bound arsenic), and F4 (crystalline iron oxide bound arsenic), according to a sequential extraction procedure using reagents of increasing dissolution strength. In this study, soil samples were collected from the vicinity of a silver smelting plant in Chenzhou, Hunan Province, and the contribution of different arsenic speciation (F1, F2, F3, and F4) to arsenic release during anaerobic enrichment incubation of paddy soil was investigated. Sample analysis was conducted at the end of the first phase (day 15) and the second phase (day 30). The effects of amorphous iron oxides in paddy soil on migration and transformation of arsenic were discussed. Results showed significant elevation of dissolved Fe(Ⅱ) and arsenic concentration (P<0.05) in enrichment solutions in the second phase compared with that in the first phase. Arsenic released in the soil solution in both phases originated from exchangeable arsenic and specifically sorbed arsenic, as indicated by its significantly positive correlation with F1 and F2 (r=0.73, P<0.05; r=0.657, P<0.05). However, an insignificant positive correlation was found between the arsenic released and F3. Moreover, HCl-extractable Fe(Ⅱ) was significantly and positively correlated with arsenic (r=0.577, P<0.05; r=0.613, P<0.05), while amorphous iron oxides were significantly and negatively correlated with arsenic (r=-0.428, P=0.126; r=-0.564, P<0.05). In conclusion, arsenic in the F1 and F2 fractions acted as the major source of released arsenic. Despite elevated levels of HCl-extractable Fe(Ⅱ) that might result from the slight reductive dissolution of amorphous iron oxide, the significant negative correlation between dissolved arsenic and amorphous iron oxides indicated that metastable amorphous iron oxides in anaerobic paddy soil can generally sorb dissolved arsenic effectively, resulting in lower mobility of arsenic. Increasing the level of amorphous iron oxides in paddy soil is conducive to inactivation of arsenic.

Preparation of Monodisperse Hydrophilic Quantum Dots with Amphiphilic Polymers.

Monodisperse hydrophilic quantum dots (QDs) are promising labeling materials for biomedical applications. However, the controllable preparation of monodisperse hydrophilic QDs with amphiphilic polymers remains a challenge. Herein, the molecular structures of amphiphilic polymers assembled on different-sized QDs are investigated. Both the experimental results and the molecular dynamics (MD) calculation suggest that the grafting ratio of amphiphilic polymers assembled on QDs increases as the size of QDs increases. Thus, the controllable preparation of different-sized monodisperse hydrophilic QDs can be achieved by simply varying the grafting ratio of amphiphilic molecules and applied in the simultaneous labeling of three tumor biomarkers.

[Influencing Mechanism of Eh, pH and Iron on the Release of Arsenic in Paddy Soil].

The massive release of soil arsenic and its enrichment in rice are significantly associated with the flooded and anaerobic management in paddy soil. Soil redox potential (Eh), pH and iron oxides exert remarkable impacts on arsenic release, which remain to be explored. In this study, long-term aerobic and anaerobic as well as intermittent aerobic incubation treatments were applied to investigate the influences of Eh, pH and iron content on arsenic release. It was found that anaerobic and flooded treatment contributed to the highest arsenic release. With decreasing Eh, significant enhancement in As(Ⅲ) and As(Ⅴ) contents in soil solution was observed. Particularly, As(Ⅲ) and As(Ⅴ) contents during the second phase increased by 1.37 and 0.99 µg·L-1compared with those in the first phase. Conversely, significant reduction in soil arsenic release (P<0.05) occurred when intermittent aerobic treatment was adopted, and the lowest level of arsenic release was observed along with the longest treatment time (6 d). The exponent relationships between arsenic and soil Eh, pH and Fe2+ content were also established, which indicated that arsenic release could be accelerated by lower pH and elevated Eh. In addition, a significant positive correlation was also found between iron(Ⅱ) content and arsenic content in soil solution. Since low Eh and elevated pH served as critical factors driving arsenic release, intermittent and aerobic water management was proved to be an effective method for the inhibition of arsenic release and uptake and accumulation of arsenic by rice.

[Effect of Different Iron Minerals on Bioaccessibility of Soil Arsenic Using in vitro Methods].

To explore the effects of different iron minerals on soil arsenic bioaccessibility, ferrihydrite, goethite and hematite were used in PBET, SBRC and IVG in-vitro experiments in this study. The relationship between arsenic bioavailability in gastric, small intestinal phases and arsenic speciation was also studied. The results showed that when 1% ferrihydrite was added, arsenic bioavailability in gastric phase was 2.22%, 5.11% and 7.43% by PBET, SBRC and IVG methods, respectively, while in the small intestinal phase it was 3.39%, 2.33% and 6.18%. At an elevated ferrihydrite dosage of 2%, significant difference in arsenic bioavailability was observed in both phases (P<0.05). According to in vitro experiments, the addition of the same amount of different iron minerals had contributed to the decrease in arsenic bioavailability to varying extents in contrast with the blank group, in the descending order of ferrihydrite(F1) > goethite(G1) > hematite(H1) (F2 > G2 > H2). Total arsenic in exchangeable (F1) and specifically sorbed (F2) state was found positively correlated with arsenic bioavailability in gastric phase by PBET, SBRC and IVG methods, the correlation coefficient of which being r=0.93, P=0.002, r=0.90, P=0.004 and r=0.89,P=0.006, respectively. It was also found that arsenic bioavailability in gastric phase was positively correlated with total arsenic in F1 and F2 states by PBET(r=0.94,P=0.001) and IVG (r=0.87,P=0.009) methods, but no significant correlation was observed by SBRC method. Additionally, three in vitro experiments showed that amorphous iron bound arsenic had significant negative correlation with arsenic bioavailability in gastric phase and small intestinal phase, except that no correlation was found in small intestinal phase by SBRC method.

Purification of quantum dot-based bioprobes via high-performance size exclusion chromatography.

Due to excellent optical properties, quantum dots (QDs) have been widely applied to sensing, labeling, and imaging. For the fabrication of QD-based bioprobes, purification is usually the crucial step. Hydrophilic octylamine grafted polyacrylic acid modified QDs (OPA-QDs) were prepared, and purified by high-performance size exclusion chromatography (HPSEC) to remove excess OPA and aggregated QDs. The percentage of suspended agglomerates of OPA-QDs in the unpurified OPA-QDs increases from 4% to 31% through a year, but the purified OPA-QDs of the same batch possess excellent colloidal stability for at least one year. Subsequently, QD-based bioprobes were fabricated by the conjugation between QDs and streptavidin (SA) or antibody (IgG), generating QD-SA and QD-IgG, respectively, which were purified via HPSEC. Finally, the resulting QD-SA and QD-IgG were adopted to detect tumour markers on slices and showed specific positive signals without nonspecific adsorption, which was contrary to the unpurified QD-IgG. Thus, the HPSEC-coupled system proposed in the current work is potent and universal for the generation of purified and monodisperse QD-based bioprobes, which is promising in the nanobiodetection field.

Intracellular self-assembly based multi-labeling of key viral components: Envelope, capsid and nucleic acids.

Envelope, capsid and nucleic acids are key viral components that are all involved in crucial events during virus infection. Thus simultaneous labeling of these key components is an indispensable prerequisite for monitoring comprehensive virus infection process and dissecting virus infection mechanism. Baculovirus was genetically tagged with biotin on its envelope protein GP64 and enhanced green fluorescent protein (EGFP) on its capsid protein VP39. Spodoptera frugiperda 9 (Sf9) cells were infected by the recombinant baculovirus and subsequently fed with streptavidin-conjugated quantum dots (SA-QDs) and cell-permeable nucleic acids dye SYTO 82. Just by genetic engineering and virus propagation, multi-labeling of envelope, capsid and nucleic acids was spontaneously accomplished during virus inherent self-assembly process, significantly simplifying the labeling process while maintaining virus infectivity. Intracellular dissociation and transportation of all the key viral components, which was barely reported previously, was real-time monitored based on the multi-labeling approach, offering opportunities for deeply understanding virus infection and developing anti-virus treatment.

LB-100 sensitizes hepatocellular carcinoma cells to the effects of sorafenib during hypoxia by activation of Smad3 phosphorylation.

Hepatocellular carcinoma (HCC) is a common cancer with poor prognosis. The multikinase inhibitor sorafenib is the only clinically proved systematic treatment for HCC. However, few patients respond to sorafenib. Hypoxic microenvironments contribute to sorafenib resistance. LB-100, a serine/threonine protein phosphatase 2A (PP2A) inhibitor was previously found to be a chemosensitizer in HCC. Here, we tested whether LB-100 could sensitize HCC to the effects of sorafenib. Intriguingly, LB-100 enhanced the effects of sorafenib in HCC cells only during hypoxic environments. LB-100 dramatically increased intracellular p-Smad3 level, which was responsible for the effect of LB-100 as a sensitizer. LB-100 downregulated Bcl-2 expression and enhanced sorafenib-induced apoptosis in HCC cells. We further proved that PP2A mediated LB-100-induced p-Smad3 overexpression. In addition, p38 mitogen-activated protein kinase pathway was activated in hypoxic conditions, and enhanced p-Smad3-dependent Bcl-2 inhibition and consequent apoptosis. In conclusion, LB-100 sensitized HCC cells to sorafenib in hypoxic environments. This effect was mediated by inactivation of PP2A, resulting in enhanced level of p-Smad3. Increased p-Smad3 downregulated Bcl-2, causing increased apoptosis of HCC cells.

Insulin-like growth factor-1 induces lymphangiogenesis and facilitates lymphatic metastasis in colorectal cancer.

AIM: To investigate the expression of insulin-like growth factor-1 (IGF-1)/insulin-like growth factor-1 receptor (IGF-1R) in colorectal cancer (CRC) tissues and to analyze their correlation with lymphangiogenesis and lymphatic metastasis. METHODS: Immunohistochemistry was used to evaluate IGF-1 and IGF-1R expression and lymphatic vessel density (LVD) in 40 CRC specimens. The correlation between IGF-1/IGF-1R and LVD was investigated. Effects of IGF-1 on migration and invasion of CRC cells were examined using transwell chamber assays. A LoVo cell xenograft model was established to further detect the role of IGF-1 in CRC lymphangiogenesis in vivo. RESULTS: Elevated IGF-1 and IGF-1R expression in CRC tissues was correlated with lymph node metastasis (r = 0.715 and 0.569, respectively, P < 0.05) and tumor TNM stage (r = 0.731 and 0.609, P < 0.05). A higher LVD was also found in CRC tissues and was correlated with lymphatic metastasis (r = 0.405, P < 0.05). A positive correlation was found between LVD and IGF-1R expression (r = 0.437, P < 0.05). Transwell assays revealed that IGF-1 increased the migration and invasion of CRC cells. In vivo mouse studies showed that IGF-1 also increased LVD in LoVo cell xenografts. CONCLUSION: IGF-1/IGF-1R signaling induces tumor-associated lymphangiogenesis and contributes to lymphatic metastasis of CRC.

[Pharmacokinetics, tissue distribution and magnetic resonance's response characterstics of folic acid-O-carboxymethyl chitosan ultrasmall superparamagnetic iron oxide nanoparticles in mice and rats].

Folic acid-O-carboxymethyl chitosan ultrasmall superparamagnetic iron oxide nanoparticles (FA-OCMCS-USPIO-NPs) are a novel molecular targeting MR contrast agent. This paper reperts the pharmacokinetics and magnetic resonance response characteristics of FA-OCMCS-USPIO-NPs in normal rats and mice, and discussed its distributing regularity in animals, providing basis for tumor targeting imaging. O-phenanthroline method was used to determine iron content in rats' plasma and mice's organs following high and low doses of nanoparticles injected through tail vein, and the blood concentration-time curve was drawn, the calculated t1/2 of two groups were greater than 7 h. The results of tissue distribution showed that only a small part of nanoparticles were swallowed by the liver and spleen, while none in the heart, lung and kidney. At the same times, the phagocytosis of nanoparticles did not change with the dose. The results of MRI showed that renal excretion occurred 4 hours after injection, and signal to noise ratio (SNR) of liver and kidney returned to normal levels 24 hours after injection. There were no nanoparticles in the lungs. So a part of nanoparticles escaped from phagocytosis of liver and spleen, and it owned lower toxicity and longer half-life. indicated its use for tumor-targeting imaging. All of these indicated its use for tumor-targeting imaging.

[Effect of imatinib at different concentrations on rat C6 glioma cell apoptosis and cell cycle].

OBJECTIVE: To investigate the effect of imatinib on rat C6 glioma cell apoptosis and cell cycle. METHODS: MTT assay was used to determine the OD value of C6 glioma cells following treatment with imatinib at different concentrations (0.156, 10 and 15 micromo/L) for 24, 48 and 72 h. The cell apoptosis was assayed by Hochest/PI staining and the cell cycle changes were analyzed by flow cytometry. RESULTS: Imatinib treatment resulted in increased number of apoptotic cells in a time- and dose-dependent manner. A 72-h treatment of the cells with imatinib at 10 and 15 micromo/L caused increased cell percentage in G(0)/G(1) phase to (68.53-/+0.83)% and (70.41-/+0.62)%, (P<0.01), decreased the percentage of G(2) phase cells to (14.48-/+0.12)% and (13.84-/+2.86)% (P<0.01), and decreased the percentage of S phase cells to (16.98-/+0.72)% and (15.78-/+2.28)%, respectively (P<0.01). CONCLUSION: Imatinib can induce apoptosis and affect the distribution of the cell cycle of C6 cells in vitro.

Antitumor effects of polysorbate-80 coated gemcitabine polybutylcyanoacrylate nanoparticles in vitro and its pharmacodynamics in vivo on C6 glioma cells of a brain tumor model.

Polybutylcyanoacrylate (PBCA) nanoparticles coated with polysorbate-80 have been extensively studied for delivery of drugs into the animal models; however, 1% polysorbate-80 coated gemcitabine PBCA nanoparticles (GCTB-PBCA-NPs) remain unknown. In this study, we investigated the inhibitory effects of brain targeted 1% polysorbate-80 coated GCTB-PBCA-NPs on C6 glioma cells in vitro and in vivo. GCTB-PBCA-NPs were prepared by emulsion polymerization and freeze drying. C6 glioma cells treated with 1% polysorbate-80 coated GCTB-PBCA-NPs showed poor growth with less cell density and increased detachment. Cell morphology was also greatly altered with nuclear vacuoles, ruptured cells and dead cells. Meanwhile, by flow cytometry, the numbers of cells treated with 1% polysorbate-80 coated GCTB-PBCA-NPs showed increase in G0/G1 phase and decreased in the S phase (P<0.01) compared with the blank control. CCK-8 assay also showed that GCTB could significantly inhibited cell proliferation in a concentration dependent manner. Finally, various preparations were injected (90 mg preparation per kg body weight) into the brain tumor model, which was produced after inoculating C6 glioma cells into Sprague Dawley (SD) rats for 14 days, it was shown that 1% polysorbate-80 coated GCTB-PBCA-NPs could significantly extend the survival time compared with the saline control (P<0.05). Taken together, 1% polysorbate-80 coated GCTB-PBCA-NPs can effectively inhibit the growth of C6 glioma cells in vitro and enhance antitumor activity on brain tumor in vivo.

[Effects of bagasse polysaccharide on the immune functions of immunosuppressed mice].

OBJECTIVE: To observe the effects of bagasse polysaccharide on the immune functions of immunosuppressed mice. METHODS: Immunosuppressed mouse models were established by intraperitoneal injections with cyclophosphamide followed by daily intragastric administration of bagasse polysaccharide. After the treatments, the mice were examined for immune organ weight index, phagocytotic function of the macrophages, delayed type hypersensitivity, serum IgM level following exposure to chicken red blood cells, formation of hemolytic plaques, T cell percentage and lymphocyte transformation. RESULTS: Treatment of the immunosuppressed mice with bagasse polysaccharide at the daily dose of 200 and 400 mg/kg significantly increased the weight of the immune organs, phagocytotic function of the macrophages, delayed type hypersensitivity, serum IgM level against chicken red blood cells, formation of hemolytic plaques, T cell percentage and lymphocyte transformation. CONCLUSION: Bagasse polysaccharide can enhance the immune functions of immunosuppressed mice.

[Island skin flap supplied by superficial temporal vessel for the whole ear lobe defect].

OBJECTIVE: To introduce a new method for reconstruction of the whole ear lobe defect. METHODS: The free island skin flap supplied by superficial temporal vessel which was designed at the area anterior and superior to crus helicis. The flap was transferred through subcutaneous tunnel and self-folded to reconstruct the whole ear lobe defect. RESULTS: Since 1999, 6 cases were treated with no complication. The ear lobe shape and skin colour were very natural. CONCLUSIONS: The island skin flap supplied by superficial temporal vessel is very suitable for the whole ear lobe defect with good cosmetic results.

[Treatment of recurrent condyloma acuminatum with solid lipid nanoparticle gel containing podophyllotoxin: a randomized double-blinded, controlled clinical trial].

OBJECTIVE: To observe the clinical efficacy and safety of podophyllotoxin delivered via solid lipid nanoparticle gel for topic treatment of recurrent condyloma acuminatum. METHODS: In a randomized double-blinded study, podophyllotoxin solid lipid nanoparticles gel and routine podophyllotoxin gel preparation was applied respectively for treatment of 97 volunteer patients with recurrent condyloma acuminatum. The therapeutic effect, condyloma acuminatum relapse following the treatment and adverse effect were evaluated. RESULTS: The wart clearance rate in the condyloma acuminatum patients in the first treatment course with podophyllotoxin solid lipid nanoparticle gel reached 97.1%, close to that with the routine preparation of 90.6%, but the nanoparticle preparation significantly reduced the recurrence rate and adverse effect (P<0.01). CONCLUSION: Podophyllotoxin delivered via solid lipid nanoparticle gel can effectively clear condyloma acuminatum and reduce its recurrence rate with only mild, tolerable adverse effect.

[Preparation and characterization of superparamagnetic iron oxide nanoparticles].

OBJECTIVE: To determine the physical and magnetic properties of superparamagnetic iron oxide (SPIO) nanoparticle prepared in our laboratory and evaluate its possibility for use as contrast agents in magnetic resonance imaging (MRI). METHODS: The SPIO nanoparticle was obtained by means of classical coprecipitation in dextran solution and its size determined by electron microscopy and photon-correlation spectroscopy. The iron content was determined by phenanthroline photometry, and T(2) values as well as relaxivity evaluated with a clinical MR system at 1.5T. RESULTS: Dextran-coated magnetite particles with a hydrodynamic diameter of 85.9 nm were prepared. The iron core size was 15 nm and the formation of Fe(3)O(4) crystal in SPIO nanoparticles was confirmed by X-ray diffraction (XRD) analysis. These particles possessed some characteristics of superparamagnetic and show a smaller spin-spin relaxation, with relaxivity and saturation magnetization of 0.1567 mmol(-1)/ms(-1) and 80 emu/g Fe, respectively. CONCLUSIONS: A stable SPIO nanoparticle with a dextran coating have been developed, and in vitro evaluation of its physical and magnetic properties suggests its potential for use as the contrast agent in MRI.

[Sampling of exhaled gas after intravenous administration of octafluoropropane-containing human albumin micropheres in dogs].

OBJECTIVE: To develop a method for quantitative collection of exhaled gas in anesthetized dogs at given time following intravenous administration of octafluoropropane (OFP)-containing human albumin micropheres for assessing the gas kinetics of OFP. METHODS: OFP-containing albumin micropheres were administered intravenously at 0.4, 0.8 and 1.2 ml/kg, respectively, in anesthetized and ventilated dogs. The exhaled air samples were analyzed by gas chromatography-tandem mass spectrometry (GC-MS-MS). RESULTS: The correlation curve between the area under curve (AUC) and administered dose was roughly linear (Y=1162.5X-417.38, r square=0.949 9). The total recovery rate was (119.49-/+27.62)% which was not significantly different from the rate of 100% (P>0.05). GC-MS-MS was accurate, sensitive, precise and applicable for OFP determination. CONCLUSION: The sampling method is useful for characterizing OFP pharmacokinetics in dogs, and also applicable for studying the pharmacokinetics of other gas-containing drugs.