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Selected AbstractsEndpoint quantitative PCR assays for Bacteroides forsythus, Porphyromonas gingivalis, and Actinobacillus actinomycetemcomitansJOURNAL OF PERIODONTAL RESEARCH, Issue 5 2003J. D. Rudney Background:, Conventional polymerase chain reaction (PCR) assays for periodontal pathogens are so sensitive that they detect infections of no clinical significance. Quantitative PCR (qPCR) may provide a solution to this problem. However, most qPCR systems require expensive real-time thermal cyclers. Objective:, Our goal was to develop qPCR assays which would allow endpoint quantification. Materials and methods:, 16S rRNA primers for Bacteroides forsythus, Porphyromonas gingivalis, and Actinobacillus actinomycetemcomitans were adapted to the AmplifluorÔ qPCR system, which incorporates fluorescein into the PCR product so that endpoint fluorescence is proportional to the original amount of template. DNA dilutions representing known numbers of cells were used as standard curves. Pooled subgingival plaques from the four deepest pockets of 21 severe adult periodontitis patients were assayed. Buccal molar supragingival plaque from 35 dental students provided healthy controls. Endpoint fluorescence was measured with a microplate reader. Results:, Optimized standard curves were linear in log,log or semilog fits over a range of 100,106 cells. Countable B. forsythus was present in all patients, with counts (as logs) from 2.4 to 7.3 (mean = 5.0), and 11 controls with counts from 2.1 to 4.5 (mean = 3.0). P. gingivalis was present in 11 patients and no controls, with counts from 2.2 to 4.7 (mean = 3.2). A. actinomycetemcomitans was present in two patients, with counts of 1.5 and 3.5. Conclusions:, AmplifluorÔ qPCR assays discriminated between plaque samples differing by one log or more, allowing major infections to be distinguished from minor ones. This approach allows high-throughput qPCR of plaque samples, using equipment available to many laboratories. [source] Evaluation of creatine transport using Caco-2 monolayers as an in vitro model for intestinal absorptionJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2001Alekha K. Dash Abstract Creatine is a nutraceutical that has gained popularity in both well-trained and casual athletes for its performance-enhancing or ergogenic properties. The major disadvantages of creatine monohydrate formulations are poor solubility and oral bioavailability. In the present study, creatine transport was examined using Caco-2 monolayers as an in vitro model for intestinal absorption. Confluent monolayers of Caco-2 cells (passage 25,35) were used for the permeability studies. Monolayers were placed in side-by-side diffusion chambers. 14C-Creatine (0.1,0.5 ,Ci/mL) was added to either the apical or basolateral side, and the transport of the creatine across the Caco-2 monolayer was measured over a 90-min period. The apical to basolateral transport of 14C-creatine was small, ranging from 0.2,3% of the original amount appearing on the receiver side in a 90-min period. Interestingly, the basolateral to apical permeability of radiolabeled creatine was substantially greater than that observed in the apical to basolateral direction. Studies with drug efflux transport inhibitors indicate that neither the P-glycoprotein nor multidrug resistance-associated protein is involved in the enhanced basolateral to apical transport of creatine. © 2001 Wiley-Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 90:1593,1598, 2001 [source] Probing the origin of the dark material on IapetusMONTHLY NOTICES OF THE ROYAL ASTRONOMICAL SOCIETY, Issue 3 2010F. Tosi ABSTRACT Among the icy satellites of Saturn, Iapetus shows a striking dichotomy between its leading and trailing hemispheres, the former being significantly darker than the latter. Thanks to the Visual and Infrared Mapping Spectrometer (VIMS) imaging spectrometer on-board Cassini, it is now possible to investigate the spectral features of the satellites in Saturn system within a wider spectral range and with an enhanced accuracy than with previously available data. In this work, we present an application of the G-mode method to the high resolution, visible and near-infrared data of Phoebe, Iapetus and Hyperion collected by Cassini/VIMS, in order to search for compositional correlations. We also present the results of a dynamical study on the efficiency of Iapetus in capturing dust grains travelling inwards in Saturn system with the aim of evaluating the viability of the Poynting,Robertson drag as the physical mechanism transferring the dark material to the satellite. The results of spectroscopic classification are used jointly with the ones of the dynamical study to describe a plausible physical scenario for the origin of Iapetus' dichotomy. Our work shows that mass transfer from the outer Saturnian system is an efficient mechanism, particularly for the range of sizes hypothesized for the particles composing the newly discovered outer ring around Saturn. Both spectral and dynamical data indicate Phoebe as the main source of the dark material. However, due to considerations on the collisional history of the Saturnian irregular satellites and to the differences in the spectral features of Phoebe and the dark material on Iapetus in the visible and ultraviolet range, we suggest a multisource scenario where now extinct prograde satellites and the disruptive impacts that generated the putative collisional families played a significant role in supplying the original amount of dark material. [source] Comparative Metabolism of ,- and ,- Peptides in the Insect Heliothis virescens and in Plant Cells of Black Mexican Sweet MaizeCHEMISTRY & BIODIVERSITY, Issue 9 2004Rob Lind The tripeptide H-Val-Ala-Leu-OH and the N -Ac-tetrapeptide amide Ac-Thr-Lys-Trp-Phe-NH2, and their , -peptidic counterparts H- ,3hVal- ,3hAla- ,3hLeu-OH and Ac- ,3hThr-(S),2hLys- ,3hTrp- ,3hPhe-NH2, respectively, have been injected into Heliothis virescens larvae and added to cell cultures of black mexican sweet maize. The body liquids of the larvae and the supernatant of the plant cell cultures were sampled 0, 2, 3, 6, 17, and/or 24,h after application and analyzed by LC/MS. While the two , -peptides were degraded rapidly in these environments, the concentration of the , -peptides was found to decrease very slowly. Thus, ca. 60% of the original amount of the , -tetrapeptide was detected in the liquids of the insect after 24,h. The plant cells did not seem to make use of the , -peptides at all, whereas, the , -tripeptide completely disappeared from the supernatant after 3,h. Thus, we have demonstrated, for the first time, the high stability of , -peptides against degradation and metabolism in an insect and a plant. Especially remarkable is the persistence of the , -tetrapeptide with its functionalised and, thus, ,metabolisable' side chains of Thr, Lys, Trp, and Phe in the insect larvae, which are known to have a high level of activity of oxidizing enzymes. The results described here match those of ADME investigations with radioactively labeled , -peptides in rats, where essentially complete stability has been observed, while environmental microorganisms have been found to biodegrade , -peptides, albeit slowly. Possible implications of these findings for biomedical and pest-control applications are proposed. [source] | ||||||||||