AIM: To establish the GC-fingerprint chromatogram of Mylabris. METHODS: Mylabris from Guizhou Province was used under the conditions of Agilent Technoligies DB-1 capillary column and the program 160℃—2℃/min—210℃(10min), and the GC fingerprint chromatogram of Mylabris was set up. RESULTS: The methodological evaluation showed that this method had a good reproducibility, common peaks' relative areas of different samples were different. CONCLUSION: This method is reliable to evaluate the quality of Mylabris.

Objective: To determine cinnamaldehyde and cinnamic acid in Ramulus Cinnamomi and Guizhifuling Capsules.Methods: RP HPLC method was used. The samples were separated on a Zobax ODS column with the mobile phase of acetonitrile:0.1% phosphoric acid (34∶76) and the detection wavelength was set at 285nm.Results: The calibration curves were linear in the range of 3.34～53.4?g?mL -1 in cinnamaldehyde ( r= 0.9996) and 0.804～12.9?g?mL -1 in cinnamic acid ( r =0.9999),respectively. The average recoveries of cinnamaldehyde in Ramulus cinnamomi and Guizhifuling capsules were 98.6% ( RSD =1.35% n=6 ) and 97.4% ( RSD =1.62% n =6), respectively and of cinnamic acid were 99.2% ( RSD =0.97% n=6 )and 100.0% (RSD =0.73%, n=6 ),respectively.Conclusion: The method is quick, acurate and suitable for the determination of Ramulus Cinnamomi and its compound Chinese medicinal preparations.

To evaluate the clinical therapeutic effect of the autologous bone marrow mesenchymal stem cells grafting integrating blood plasma transplantation in treatment of problematic nonunion. In March 2000, a 29-year-old man presented complaints of painful walking of left thigh after a velocity injury, was selected. Radiography revealed a left femoral shaft fracture, and the bone defect distance was 5 mm through X-ray examination before operation. Following a series of reposition, fixation, intramedullary nail fixation and twice autogenous lilac bone graft treatment, external fixator and autogenous lilac bone graft treatment, totally four times, the fracture was not healed. He come to Orthopaedic Surgery Department of Siping Central Hospital to accept the autologous bone marrow mesenchymal stem cells grafting integrating blood plasma transplantation in March 2008. Under the small C-arm X-ray perspective, 4 mL bone marrow-derived mesenchymal stem cells suspension integrating 10% autologous blood plasma was injected vertically into fracture site from the skin in front of the thigh with epidural needle, and the stem cells density was 1.8 × 107 cells/L. X-ray examination was performed though out-patient recheck to observe fracture healing. Two months after graft, radiography results showed fracture interspace reduced, left femoral shaft callus was continual and fracture lines blurred partly of the left femur; furthermore, left femoral shaft fracture lines blurred, and continuous bone callus formation of the left femur at 4 months after graft; The left femur achieved bone union within 7 months. The patient was returned to full weight bearing walking and good function with a fully healed femoral bone, without any fever or infection. Percutaneous transplantation of autologous bone marrow stem cells for treatment of problematic nonunion has the satisfactory result.

AIM: To establish chromatographic fingerprint of Panax gingseng C.A.Mey.by HPLC. METHODS: Separation was performed on an Agilent ZORBAX SB-C_(18)(4.6 mm?250 mm,5 ?m) analytical column with mobile phase consisting of acetonitrile and water with gradient elution with the flow rate 1.0 mL/min and the column temperature at 40 ℃.The UV wavelength used for detection was set at 203 nm and the analysis time was 90 min. RESULTS: 18 co-peaks on the HPLC fingerprints of Panax ginerseng C.A.Mey.were indicated.The similarities were determined by cosine.The results of similarity analysis were 0.90～1.00. CONCLUSION: Perfect fingerprints were obtained which can be used for the quality control of Panax gingseng C.A.Mey.

In order to construct the expression recombinant of human receptor associated protein (RAP), optimize its expression condition and obtain the recombinant protein after expression with high efficiency, two prokaryotic expression vectors-pT7-PL and pET-28a(+) were used to construct the expression recombinant containing RAP cDNA, and the expression efficiency of two kinds of expression E. coli of BL21 strains was compared. The effect of different induction conditions on the expression of recombinant RAP was observed. After recombinant protein was purified with Ni(+) -nitrilotriacetic acid (Ni(+) -NTA) affinity chromatogram, its binding ability with microphage was observed. The results showed that two recombinant plasmids both obtained high expression of RAP. The expression levels of RAP in plasmid pT7-PL-RAP in BL21 (DE3, plysS) strain were significantly higher than in BL21 (DE3) strain. The expression of pT7-PL-RAP in the presence of chloramphenicol was higher than in the absence of chloramphenicol, and most of the inducible expressed RAP was soluble. The RAP which was purified by Ni(+) -NTA resin could strongly bind with the RAW264.7 cells rich in low density lipoprotein receptor (LDLR) family receptors. It was concluded that the expression condition of recombinant RAP was optimized and functional RAP was obtained, which offered a good foundation for the further production of RAP as research tool.

A high-performance liquid chromatography coupled ultraviolet （HPLC-UV） method was developed for a chemical fingerprint analysis of Viscum coloratura. Eighteen peaks were selected as the common peaks and Homoeriodictyol-7-O-β-D-apiosiyl-（1→2）-β-D-glucoside was used as a reference. The relative areas of common peaks were used for hierarchical clustering analysis and similarity calculation. Thirty-seven samples collected from different sources were classified into five groups. The similarities of 21 batches Viscum coloratura samples were beyond 0.90. The results obtained suggest that the chromatographic fingerprint can efficiently identify Viscum coloratum. Additionally, the fingerprints can then be used to evaluate the correlation between Viscum coloratura and hosts.

A high-performance liquid chromatography coupled ultraviolet (HPLC-UV) method was developed for a chemical fingerprint analysis ofViscum coloratum. Eighteen peaks were selected as the common peaks and Homoeriodictyol-7-O-β-D-apiosiyl-(1→2)-β-D-glucoside was used as a reference.The relative areas of common peaks were used for hierarchical clustering analysis and similarity calculation.Thirty-seven samples collected from different sources were classified imo five groups.The similarities of 21 batches Viscum coloratuma samples were beyond 0.90.The results obtained suggest that the chromatographic fingerprint can efficiently identify Viscum coloratum.Additionally,the fingerprints can then be used to evaluate the correlation between Viscum coloratum and hosts.

In order to obtain three isoforms of apolipoprotein E (apoE), the cDNA encoding apoE3 was obtained by RT-PCR from normal human liver tissue. Site-directed mutagenesis was used to obtain the cDNAs encoding apoE2 and apoE4 isoforms. The 3 cDNAs were subcloned into vector pGEM-3Z and verified by DNA sequencing. The expression recombinant which can express the target protein as a (His) 6-tagged fusion was constructed by subcloning apoE cDNA into vector pT7-PL. The purified proteins were gained by Ni-NTA column. The SDS-PAGE results revealed the 6 His fusion proteins (apoE2, apoE3 and apoE4) were correctly expressed and purified successfully.

In order to construct the expression recombinant of human receptor associated protein (RAP), optimize its expression condition and obtain the recombinant protein after expression with high efficiency, two prokaryotic expression vectors-pT7-PL and pET-28a(+) were used to construct the expression recombinant containing RAP cDNA, and the expression efficiency of two kinds of expression E. coli of BL21 strains was compared. The effect of different induction conditions on the expression of recombinant RAP was observed. After recombinant protein was purified with Ni+-nitrilotriacetic acid (Ni+-NTA) affinity chromatogram, its binding ability with microphage was observed. The results showed that two recombinant plasmids both obtained high expression of RAP. The expression levels of RAP in plasmid pT7-PL-RAP in BL21 (DE3, plysS) strain were significantly higher than in BL21 (DE3) strain. The expression of pT7-PL-RAP in the presence of chloramphenicol was higher than in the absence of chloramphenicol, and most of the inducible expressed RAP was soluble. The RAP which was purified by Ni+-NTA resin could strongly bind with the RAW264.7 cells rich in low density lipoprotein receptor (LDLR) family receptors. It was concluded that the expression condition of recombinant RAP was optimized and functional RAP was obtained, which offered a good foundation for the further production of RAP as research tool.

OBJECTIVETo develop a reversed-phase HPLC method for simultaneous determination of gastrodin, adenosin, 4-hydroxybenzyl alcohol and 4-hydroxybenzaldehyde in Rhizoma Gastrodia.

METHODA Kromasil C18 column (4.6 mm x 250 mm, 5 microm) was used with a methanol-water-0.1% acetic acid gradient elution system. The eluates were detected at 270 nm, the flow rate was 1.0 mL x min(-1) and the column temperature was 35 degrees C.

RESULTThe linear range of gastrodin, adenosin, 4-hydroxybenzyl alcohol and 4-hydroxybenzaldehyde were 19.1-383 (r = 0.999 9), 0.620-12.4 (r = 0.999 9), 2.45-49.0 (r = 0.999 9), 0.280-5.63 mg x L(-1) (r = 0.999 6), respectively. The average recoveries (n = 9) of the four components were 96.7% -97.7%, RSD < 1.6%.

CONCLUSIONThe method is accurate, sensitive and reliable for determination of gastrodin, adenosin, 4-hydroxybenzyl alcohol and 4-hydroxybenzaldehyde in Rhizoma Gastrodia.