Objective To investigate the dynamic variation of flavonoids contents in the flowers of Citrus grandis at different flowering periods. Methods The content of total flavonoids in the flowers of Citrus grandis was determined by means of ultraviolet-visible absorption spectroscopy ,and the contents of naringin and rhoifolin were determined by HPLC. Results The content of total flavonoids in the flowers during the flower withering period ,the young fruit period ,the blossom period,and the budding period was 306.90 mg.g-1,277.93 mg.g-1,215.55 mg.g-1 and 162.74 mg.g-1 respectively. The naringin content during the above four different periods was 246.31 mg.g-1,213.93 mg.g-1,175.94 mg.g-1 and 130.90 mg.g-1 respectively. The rhoifolin content during the four periods was unchanged. Conclusion The contents of total flavonoids and naringin in flowers of Citrus grandis during flower withering period are the highest.

Objective To observe the effect of apigenin on the recovery of neurological function following cerebral ischemia-reperfusion and investigate its mechanism. Methods Ninety male Sprague-Dawley rats were randomized into a sham-operated group, a model group and an apigenin-treated group. A transient ( 1.5 h) focal cerebral ischemia-reperfu-sion model was established in the rats of the model and apigenin-treated groups. In the sham-operated rats the middle cere-bral artery was not occluded. The rats in the apigenin-treated group received an intra-abdominal injection of apigenin, and the rats in the other two groups received injections of normal saline solution. Neurological behavior scores were assessed in accordance with the Zea Longa method at the 24th, 48th and 72nd hour and the 7th day after reperfusion. Cellular and sub-cellular morphology were observed with an optical microscope and an electron microscope, and the levels of TNF-α and IL-1β were measured using ELISA. Results Neurological function improved by the 7th day after reperfusion in the model group, but improved significantly by the 72nd hour after reperfusion in the apigenin-treated group. Average TNF-α and IL-1β levels in the model group and the apigenin-treated group were significantly higher than in the sham-operated group. Av-erage TNF-α and IL-1β levels in the apigenin-treated group were significantly lower than in the model group at the 48th and 72nd hour after reperfusion. Neurological behavior scores had a positive correlation with the IL-1β and TNF-α levels. In the model group, obvious intracellular and intercellular edema and vacuolization were observed in the ischemic cortices and hippocampuses, with remarkable karyopycnosis and organelle broadening and dissolution and vacuolization in glial cells and neurons. In the apigenin-treated group, similar but significantly milder morphological changes were observed. Conclusion Apigenin can promote the recovery of neurological function in rats by downregulating the expression of TNF-α and IL-1βfollowing focal cerebral ischemia-reperfusion.

Objective To investigate the influence of producing areas and sexual reproduction on the genetic features of Citrus grandis‘Tomentosa’. Methods The genetic distance in different species was counted,and the DNA fingerprint of germplasm resources of Citrus grandis ‘Tomentosa’ was established after random amplified polymorphic DNA analysis of Citrus grandis ‘Tomentosa’ and Citrus grandis (L.) Osbeck of various species from different producing areas. Results The genetic distance index D of Citrus grandis ‘Tomentosa’ and Citrus grandis (L.) Osbeck of various species from different producing areas was in the range of 0.01~0.64. Conclusion Sexual reproduction and the changes of producing areas can result in the genetic diversity of Citrus grandis ‘Tomentosa’.

Objective To establish the optimal extracted method for content dete rmination of naringin in Citrus grandis. Methods RP-HPLC was used to determinat e the content of naringin extracted with the above two methods from different ye ar samples of Citrus grandis. Results The average content extracted with ultraso nic extraction was 13.53 %,and the average content extracted with soxhlet extr action was 11.98 %,there being insignificant difference between the two method s. Conclusion The content of naringin extracted with ultrasonic extraction is mo re than that with the soxhlet extraction,which be receipted in Chinese pharmeco pia. And ultrasonic extraction method is more convenient and can save time.

【Objective】 To explore the effect of different extraction methods on the content of total flavones from propolis.【Methods】With rutin as the control,spectrophotometry was used to investigate the content of total flavones extracted from propolis.【Results】The content of total flavones extracted by pharmacopoeia extraction method was 0.1982mg/g and 0.1978mg/g by methanol Soxhlet extraction method,the difference being insignificant.【Conclusion】Methanol Soxhlet extraction method is simple,effective and practical,and can be used to replace pharmacopoeia extraction method for the determination of total flavones from propolis when necessary.

Objective:To study the mechanisms that acupuncture therapy of "Shuanggu Yitong" in treating postmenopausal osteoporosis(PMO).Methods:60female SD rats aged 6 months were resected bilateral ovaries and feeded for 90 days in order to make the model of experimental osteoporosis.Then,they were divided into 5 groups such as control group,regular acupoints treated group,"Shuanggu" group,"Yitong" group,"Shuanggu Yitong" group,with 12 rats each group,and the control group without any treatment;Pishu(BL 20),Weishu(BL 21),Shenshu(BL 23),Qihaishu(BL 24) was selected for the regular acupoints treated group;Guanyuan(RN 4),Zusanli(ST 36) for the "Shuanggu" group;and Shenshu(BL 23),Geshu(BL 17),Dazhu(BL 11) for the "Yitong" group;Guanyuan(RN 4),Zusanli(ST 36),Shenshu(BL 23),Geshu(BL 17),Dazhu(BL 11)for the "Shuanggu Yitong" group.Electroacupuncture therapy was given once a day,15 minutes each time,the needles were retained for 10 minutes.Then these rats rested for 1 day after consecutive therapy for 6 days.The therapy lasted for 12 weeks.RT-PCR was used to examine the expression level of interleukin-6(IL-6)mRNA in bone tissue.Results:Compared with control group,the expression level of IL-6mRNA in all acupuncture therapy groups decreased(P0.05).Conclusion:Acupuncture can reduce the expression level of IL-6mRNA in osteoporosis model rats without ovaries and "Shuanggu Yitong" acupuncture therapy is superior to other regular acupuncture therapy.

Objective To investigate the mechanism of contrast-induced nephropathy (CIN) caused by Iopromide.Methods Two-four female SD rats were randomly divided into two groups which were control group and CIN group .The rats in CIN group were injected Io-promide via caudal vein ,the rats in control group were injected the equal amount of solvent .After 24 hours,all the rats were euthanized and tested.The excretion of 24 h urinary protein was detected using biochemistry assay .The expression of related cell cycle regulatory protein such as P21,P27 and TGF-β1 in glomerular visceral epithelium cells were measured using immunohistochemical technique .A semiquantitative score was used to evaluate the injure degree of glomerular and tubulointerstitium .Renal glomerular cell apoptosis was evaluated by TUNEL . Results Compared with control group ,CIN group rat glomerular epithelial cells of P 21,P27 and TGF beta 1 positive expression rate signifi-cantly increased,[(12.86 ±0.98) %vs (0.46 ±0.21)%,P=0.004 5],[(21.76 ±2.75)% vs (9.57 ±1.86)%,P =0.0071], [(12.85 ±5.54) vs (7.63 ±0.84),P=0.003 7)] respectively,24 h urine protein significantly increase [(23.44 ±5.22) mg/d vs (2.13 ±0.52) mg/d,P=0.007 0,P=0.005 0],CIN pathological damage of rat glomerular epithelial cells and apoptotic rate significantly more serious [(52.5 ±6.4)%vs (4.2 ±0.3) %,P =0.007 5].In addition,the renal pathologic scores were positively correlated with the excretion of 24hr urinary protein and the expression of P 21,P27,and TGF-β1(r=0.765,0.701,0.842,0.651,P<0.01).Conclusion Io-dine amine via increased glomerular epithelial cells P 27 and TGF-beta 1expression and urinary protein excretion , aggravating pathological damage and apoptosis .

Objective To set up a new diagnostic platform based on microarray exon-capture and next-generation sequencing for detecting small mutations in dystrophin gene.The sensitivity and specificity of the method were assessed in clinical settings and the distribution of small mutations in Chinese Duchenne muscular dystrophy/Becker muscular dystrophy (DMD/BMD) patients were also analyzed.Methods Forty-one DMD/BMD patients diagnosed by the clinical criteria without large deletion or duplication (≥ 1exon) were recruited from Peking Union Medical College Hospital consecutively.Genomic DNA was extracted from blood samples.The libraries were prepared.Then exon and intron-exon flanking sequences of DMD gene were captured by custom microarray.Targeted next-generation sequencing and Sanger Sequencing were conducted.The patients who were not detected any disease-causing mutation were performed muscle biopsy.Results Thirty-eight subjects were detected small mutations in DMD gene.All single nucleotide variants (SNVs) and insertion & deletions (INDELs) were validated by Sanger sequencing.Twenty-one novel mutations were reported.The distribution of SNVs and INDELs was similar to other international DMD databases.Upon immunohistochemistry staining of dystrophin protein,1 of 3 mutation-undetected patients was diagnosed as DMD,2 of them were excluded.The specificity of the method was 100％,while the sensitivity was 97.4％.Conclusions Our microarray-captured next-generation sequencing assay could detect SNVs and INDELs with high sensitivity and specificity.Its advantages are economic,time-saving and stable.The platform is suitable for clinical gene diagnosis.

Objective To establish an early cognitive disorder model in rats and investigate the early cognitive functioning after ischemic hypoxic brain injury during the neonatal period. Methods Forty-six newborn Sprague-Dawley rats were randomized into a 21-d-old group and a 31-d-old group. These 2 groups were then subdivided into model and sham-operated subgroups (M21, n=12; SH21, n=11; M31, n=12; SH31, n=11). A model of neonatal early cognitive disorder was established in the rats of the M21 and M31 groups using a modification of Rice's method. Rats in the SH21 and SH31 groups received skin incisions and common carotid artery separation without ligation or hypoxia. Each group was tested with a Morris water maze. The rats were sacrificed after testing, and brain tissue was examined under the electron microscope. Nissl staining allowed Nissl body quantification and neurocyte acin the M21 group was significantly longer than in the SH21 group. The 31-d-old subgroups had shorter average escaping latencies than the corresponding 21-d-old subgroups. (b) Spatial memory: The average platform times, Ⅰ region times and Ⅰ region distances showed no significant differences among groups. ②Brain pathology (a) Gross appearance: Obvious ischemic hemisphere atrophy was observed in the M group, and no abnormality was observed in the SH group. (b) Electron microscopic observation: In the SH group cell ultrastructures in the ischemic hippocampus were normal. Karyopyknosis and dilated endoplasmic reticulums were found in the M group. More mitochondria were found in the presynaptic membranes of the ischemic hippocampus in the M group than that in the SH group. (c) Nissl body quantification and neurocyte activity analysis: Significantly less activity in the ischemic cortex was found in the M21 group compared to the SH21 group. More activity was observed in the 31-d-old subgroups than in the corresponding 21-d-old subgroups. Conclusions ①The neonatal rats with ischemic hypoxic brain injury had prolonged average escaping latency and depressed neuronal activity. ②The 31-d-old rats had better spatial localization learning ability than the 21-d-old rats.

Objective To explore the genuineness of Exocarpium Citri Grandis(ECG) from Huazhou city of Guangdong province.Methods We used the method of high performance liquid chromatography to detect the naringin content in ECG from different producing areas of Huazhou city.Random amplified polymorphic DNA analysis was used for the examination of genetic distance,and plasma-atomic emission spectrometry for the detection of soil elemental abundance of 8 elements such as aluminium(Al),kalium(K),calcium(Ca),ferrum(Fe),titanium(Ti),boron(B),magnesium(Mg),and manganese(Ma).The correlation of the above three parameters was analyzed by statistical software SPSS 11.5.Results Ca abundance in the surface soil layer had an obvious effect on the content of naringin,and the difference of Al and K abundance in subsoil layer was correlated with the genetic distance of ECG.Conclusion The genuineness of ECG is probably related with the abundance of phlopopitum in the soil of producing areas of Huazhou city.