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Objective: To determine the content of index components in different parts of Gardenia jasminoides (pericarp, seeds, whiskers), study the fingerprint, and compare the contents and compositions differences of different parts of G. jasminoides, in order to provide the theoretical basis for different efficacies of G. jasminoides pericarp and seeds, explore the exploitation and utilization values of G. jasminoides whiskers, and avoid waste of gardenia medicinal resources. Method: The contents of geniposide and crocetin Ⅰ was were determined by HPLC, the content of total iridoid glycosides was determined by ultraviolet spectrophotometry, and three index components in different parts of G. jasminoides were analyzed. HPLC fingerprints of different parts of G. jasminoides were collected, the common pattern of HPLC fingerprints of different parts of G. jasminoides of different origins and with different processing methods was established, and the similarity evaluation software was used for data analysis; comparative analysis on fingerprints of different parts of G. jasminoides was conducted. Result: Content change of index components in G. jasminoides pericarp and seeds from Henan, Fujian and Jiangxi were the same. Content of geniposide:Fujian > Henan > Jiangxi, the contents of three components in G. jasminoides pericarp from Fujian were much higher than those from Henan and Jiangxi, the contents of crocetin Ⅰ and total iridoid glycosides:Fujian > Jiangxi > Henan, the contents of total iridoid glycosides from Fujian, Jiangxi were much higher than those from Henan. The order of three index components in G. jasminoides whiskers from different origins from high to low, the content of geniposide and crocetin Ⅰ was Fujian > Jiangxi and Henan, the content of total iridoid glycosides was Fujian > Jiangxi > Henan.In the same part, there were 22 common peaks in the fingerprints of G. jasminoides pericarp, except for S13-S15, the similarity of other samples were more than 0.9;the fingerprints of G. jasminoides seeds had 22 common peaks, except for S22-S30, the similarities of other samples were more than 0.9;the fingerprints of G. jasminoides whiskers had 16 common peaks, except for S7-S9, the similarities of other samples were more than 0.9.In different parts, the fingerprints of G. jasminoides whiskers were significant different from those of pericarp and seeds, the number of peaks in G. jasminoides whiskers reduced, the order of height of peaks 2, 3, 5 of G. jasminoides from high to low were whiskers > gardenia > seeds. There was not peak X in the seeds, the height of peak X of gardenia in whiskers was higher than that in pericarp, except for the peak 17, the height of all peaks in seeds were higher than that in whiskers. Conclusion: There are significant differences in the contents of index components in G. jasminoides pericarp and seeds. The content of total glycosides in gardenia is high, suggesting that it can be used to extract total iridoid glycosides. The fingerprints can reflect the content difference and species distribution of different parts of G. jasminoides, so as to provide theoretical support for the studies for pharmacodynamic material basis of G. jasminoides and the scientificity and rationality of the separate application of G. jasminoides pericarp and seeds.
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Objective:To study HPLC fingerprints of Achyranthis Bidentatae Radix from different origins,compare different specifications in the same origin,and explore the effect of origin and specifications on the quality of Achyranthis Bidentatae Radix and relationship between the specifications and the internal quality of Achyranthis Bidentatae Radix, in order to provide basis for the identification of its origin. Method:The HPLC fingerprints of Achyranthis Bidentatae Radix from different origins and with different specifications in the same origin were collected. The similarity analysis,cluster analysis and principal component analysis were adopted to analyze the fingerprints,the differences in fingerprints of Achyranthis Bidentatae Radix from different origins and with different specifications in the same origin were compared. Result:Analysis of different origins and principal component analysis could be used to distinguish Achyranthis Bidentatae Radix from five producing areas,and the identification results of origin analysis was better than those of cluster analysis and similarity analysis. Analysis of different specifications, similarity analysis or principal component analysis could not distinguish Achyranthis Bidentatae Radix with different specifications. Conclusion:There are significant differences in chemical composition and peak height among Achyranthis Bidentatae Radix from different origins,with less differences in chemical composition and peak height of Achyranthis Bidentatae Radix with different specifications, the principal component analysis could be used to identify origins of Achyranthis Bidentatae Radix.
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Objective:To investigate and compare the fingerprints of different polarity fractions (petroleumether,chloroform,ethyl acetate,n-butanol,water) of fresh Gardeniae Fructus with different fruit shapes,and further understand the content difference and distribution of its chemical composition. Method:Gardeniae Fructus was reflux extracted by water in order to obtain the water extract; water extract 0.1 g and dissolved with 50 mL water,then it was extracted by petroleum ether,chloroform,ethyl acetate and n-butanol in turn in order to obtain the different extraction phases and the water phase. Each phase was condensed to extractum. Finally,the samples were analyzed by high performance liquid chromatography (HPLC) fingerprints and the similarity evaluation software was used for data analysis. Result:Fingerprint of chloroform fraction of water extract in gardenia from different habitats can be used to distinguish Gardeniae Fructus from Fujian,Henan and Jiangxi. The differences between the water extract of Gardeniae Fructus from Fujian and those of Henan and Jiangxi were mainly manifested in the petroleum ether fraction, and the fat-soluble components of Gardeniae Fructus were more than those of Henan and Jiangxi. The differences between the water extract of Gardeniae Fructus from Henan and those of Fujian and Jiangxi were mainly manifested in the ethyl acetate fraction,and the content of iridoid glycosides was significantly higher than that in Fujian and Henan. The differences between the water extract of gardenia from Jiangxi and those of Fujian and Henan were mainly manifested in the n-butanol fraction,organic acid peak C1 not detected. The fingerprint of chloroform fraction of water extract in Gardeniae Fructus can be used to distinguish Gardeniae Fructus of six ribs and Gardeniae Fructus of seven ribs from Fujian and Henan,and the contents of all components of Gardeniae Fructus of seven ribs were more than those in Gardeniae Fructus of six ribs. The fingerprint of petroleum ether fraction of water extract in Gardeniae Fructus can be used to distinguish Gardeniae Fructus of six ribs and Gardeniae Fructus of seven ribs from Jiangxi. The Z3 peak of Gardeniae Fructus of six ribs from Henan was obviously higher than that of Gardeniae Fructus of seven ribs. The contents of all components on chloroform and ethyl acetate fractions of water extract in Gardeniae Fructus of seven ribs were significantly higher than those of Gardeniae Fructus of six ribs. Conclusion:There are significant differences on chemical constituents and content among Gardeniae Fructus from Fujian,Henan and Jiangxi. The main difference of fingerprint between Gardeniae Fructus of six ribs and Gardeniae Fructus of seven ribs is the peak height.
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Objective To determine the content of baicalin in Radix Scutellariae granule from different manufacturers using near infrared spectroscopy(NIRS)technology. Methods Utilizing NIRS combined with partial least squares, and simultaneously optimizing the pretreatment methods and the range of spectrum, the quantitative model of baicalin in Radix Scutellarie granules was established. Results In the model correlation coefficient(R2)was 0.9702, root-mean-square error of calibration(RMSEC)was 0.555, and root-mean-square error of predict(RMSEP)was 1.05. Conclusion In this paper, rapid quality analysis method of Radix Scutellariae granules from different manufacturers is studied using NIRS technology. The model can predict the content of baicalin nondestructively and rapidly. It can give some reference for researching the quality control of Radix Scutellariae granules.
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<p><b>BACKGROUND</b>Elevated intraocular pressure (IOP) is primarily due to increased aqueous outflow resistance, but how aqueous outflow resistance is generated and regulated are still not fully understood. The aim of this study is to determine whether changes in outflow facility, outflow pattern, and morphology following acute IOP elevation were reversible when the IOP was returned to a normal level in bovine eyes using a two-color tracer technique to label outflow patterns within the same eye.</p><p><b>METHODS</b>Twelve fresh enucleated bovine eyes were perfused with Dulbecco's phosphate buffer saline (PBS) containing 5.5 mmol/L glucose (DBG) at 30 mmHg first to establish the baseline outflow facility followed by a fixed volume of red fluorescent microspheres (0.5 µm, 0.002% v/v). After the red tracer being replaced with DBG in the anterior chamber, perfusion was continued at 7 mmHg with the same volume of green tracer, followed by a fixative. In two control groups, the eyes were constantly perfused at either 30 mmHg (n = 6) or 7 mmHg (n = 6) using the same methods. The outflow facility (C, µl × min × (-1)mmHg(-1)), was continuously recorded. Confocal images were taken along the inner wall (IW) of the aqueous plexus (AP) in frontal sections. The percent of the effective filtration length (PEFL, PEFL = IW length exhibiting tracer labeling/total length of IW) was measured. Sections with AP were processed and examined by light microscopy. The total length of IW and the length exhibiting separation (SL) in the juxtacanalicular connective tissue (JCT) were measured. A minimum of eight collector channel (CC) ostia per eye were analyzed for herniations.</p><p><b>RESULTS</b>In the experimental (30 - 7 mmHg) group, the outflow facility was significantly higher at 7 mmHg ((4.81 - 1.33) µl × min × (-1)mmHg(-1)) than that at 30 mmHg ((0.99 ± 0.15) µl × min × (-1)mmHg(-1), P = 0.002), corresponding to a significant increase in the PEFL (P = 0.0003). The percent of CC ostia exhibiting herniations in the experimental group ((67.40 ± 8.90) µl × min × (-1)mmHg(-1)) decreased significantly compared to that in the control at 30 mmHg ((94.44 ± 3.33) µl × min × (-1)mmHg(-1), P = 0.03), but higher than that in the control at 7 mmHg ((29.43 ± 4.60) µl × min × (-1)mmHg(-1), P = 0.01). Washout-associated separation between the IW and JCT was found by light microscopy and percent separation length (PSL, PSL = SL/total length of IW) was decreased in the control at 30 mmHg compared to that in the experimental group and control at 7 mmHg.</p><p><b>CONCLUSIONS</b>The pressure-induced morphological and hydrodynamic changes were reversible. Changes (collapse of AP, separation between the JCT and IW, and herniation into CC ostia) influence the effective filtration area that regulates outflow facility.</p>
Subject(s)
Animals , Cattle , Aqueous Humor , Physiology , Hydrodynamics , Intraocular Pressure , Physiology , Microscopy, ConfocalABSTRACT
<p><b>BACKGROUND</b>Elevation of intraocular pressure is usually associated with primary open angle glaucoma and caused by increased outflow resistance. A two-color fluorescent tracer technique was developed to investigate the hydrodynamics of aqueous humor outflow with changing intraocular pressure within the same eye, to better understand the relationship between outflow facility and effective filtration area.</p><p><b>METHODS</b>Eighteen enucleated bovine eyes were first perfused at 30 mmHg with Dulbecco's phosphate-buffered saline containing 5.5 mmol/L D-glucose. After a stable baseline facility, red fluorescent microspheres (0.5 microm, 0.002% v/v) were exchanged and perfused. Eyes in the one-color control group (n = 6) were immediately perfused with fixative. In the experimental group (n = 6), eyes were perfused with green tracer after intraocular pressure reduced to 7 mmHg, while in the two-color control group (n = 6), eyes were perfused with green tracer with intraocular pressure remaining at 30 mmHg. All 12 eyes were then perfusion-fixed. Outflow facility was continuously recorded in all eyes. Confocal images were taken along the inner wall of the aqueous plexus and the percent of the effective filtration length (PEFL; length of inner wall exhibiting tracer labeling/total length of inner wall) was measured. The relationships between outflow facility and PEFL were analyzed statistically.</p><p><b>RESULTS</b>No significant differences were found in baseline facilities (microl x min(-1) x mmHg(-1)) among the three groups (the experimental group: 0.93 +/- 0.12; the two-color control group: 0.90 +/- 0.19; the one-color control group: 0.98 +/- 0.13). In the experimental group, the outflow facility was significantly higher at 7 mmHg (4.29 +/- 1.01) than that at 30 mmHg (1.90 +/- 0.67, P < 0.001), which corresponded to a significant increase in the PEFL at 7 mmHg (54.70 +/- 8.42) from that at 30 mmHg ((11.76 +/- 4.56)%, P < 0.001). The PEFL labeled by red fluorescent microspheres in the experimental group ((11.76 +/- 4.56)%) showed no significant difference from that of the one-color control group ((13.39 +/- 2.19)%, P = 0.473) or the two-color control group ((11.49 +/- 4.95)%, P = 0.930). The PEFL labeled by green fluorescent microspheres in the experimental group ((54.70 +/- 8.42)%) was significantly higher than that of the two color control group ((37.34 +/- 8.17)%, P = 0.010). A positive correlation was found between outflow facility and PEFL (r = 0.897, R(2) = 0.804) in the experimental group.</p><p><b>CONCLUSIONS</b>Changes in aqueous humor outflow patterns before and after a change in intraocular pressure can be successfully distinguished within the same eye using our newly developed two-color tracer perfusion technique. The PEFL showed positive correlation with the outflow facility.</p>
Subject(s)
Animals , Cattle , Aqueous Humor , Physiology , Intraocular Pressure , Luminescent Proteins , Metabolism , Microscopy, Confocal , Microspheres , Perfusion , MethodsABSTRACT
Ticks are obligate ectoparasites and vectors of arboviruses, vickettsiate, spirochetes and parasitil protozoa of humans and domestic animals. Immunological protection of mammalian hosts against tick infestation has been proposed as the most sustainable alternative tick control method to the current use of acaricides. The success of this method is dependent on the identification of key molecules for use as tick vaccine antigens. Proteolytic enzymes are involved in a wide range of cellular processes, thus they can be considered as good target antigens for a tick vaccine. In the present study, we used rapid amplification of cDNA ends protocol and primers that were designed based on the consensus amino acid motifs flanking present in all papain-like cysteine proteinases, to amplify, sequence and characterize two Rhipicephalus haemaphysaloides haemaphysaloides cathepsin L-like cysteine proteinases, named as cysA and cysB. The full length of cysA is 1168bp, encoding a 332 amino acid residue polypeptide with 36.33kD predicted molecular mass; the full length of cysB is 1153bp, encoding a 335 amino acid residue polypeptide with 37.56kD predicted molecular mass. The consensus amino acid motifs flanking presence in both deduced amino acid sequences. And both genes show high sequence homology to other tick cathepsin L-like cysteine proteinase, so they were identified as members of the cysteine proteinase gene family. Expression analysis by RT-PCR revealed that cysA and cysB were expressed differently in different periods of tick development.