Simple high-performance liquid chromatography method for the simultaneous determination of serum caffeine and paraxanthine following rapid sample preparation

A simple reversed-phase high-performance liquid chromatography (HPLC) method for the simultaneous determination of caffeine and paraxanthine in human serum is described. Serum proteins are precipitated with perchloric acid and the resulting supernatant neutralized for direct injection onto an HPLC column. The method uses a phosphate–methanol mobile phase (85:15, v/v) at pH 4.9 with a flow-rate of 1.75 ml/min and quantitation is by UV absorbance at 274 nm. Elution times are approximately 18 min for caffeine and 8 min for paraxanthine. Theobromine and theophylline have elution times of 5.4 and 9.4 min and do not interfere in the assay. The intra-assay and between-assay means for precision and accuracy for both drugs are: 4.5% C.V. and 3.3% deviation. The sensitivity of the method is 50 ng/ml for each drug.     

Comparison of antiprogestin stimulation of uterine prostaglandin synthesis in vitro

Progesterone has an inhibitory effect on prostaglandin synthesis in urine tissue and this effect is reversible with progesterone receptor antagonists. Although antiprogesterone steroids such as RU486 (Mifepristone) are effective at inducing abortion in women they have an improved efficacy when used with exogenous synthetic prostaglandin. In the guinea-pig such antagonists sensitize the uterus but do not result in increased myometrial activity and therefore may not induce endogenous PG synthesis. In this study the effects of antiprogestins on a preparation of rat uterus perifused with progesterone were studied. ZK98 734 caused a rapid and sustained increase in 6-oxoPGF synthesis which rose within the first 90 minutes. This rapid response suggested that some mechanism other than the induction of fresh protein synthesis was involved. A similar increase was not seen with pregnant guinea-pig myometrium/decidua perifused in a similar manner, suggesting that some other mechanism was responsible for the relatively low PG production in pregnancy. However increases in 6-oxoPGF in response to antiprogestins were recorded when pregnant guinea-pig decidua/myometrium was incubated for 4 hours. In these experiments 1 microM ZK98 734 and 1 microM ZK98 299 (Onapristone) gave a 2.7 fold increase in PG production whereas RU486 gave a 1.6 fold increase. Both 1 microM ZK98 734 and 1 microM ZK98 299 also gave a significant increase in PGE production but no increase in PGF was observed. These findings suggest that some antiprogestins might have a better effect on the stimulation of endogenous PG synthesis or on the rate of catabolism of prostanoids.     

Fertilization acid of sea urchin eggs: Evidence that it is H+, not CO2

The report of R. J. Gillies, M. P. Rosenberg, and D. W. Deamer (1981, J. Cell. Phys., 108, 115–122) that sea urchin fertilization acid is anaerobically produced CO₂, was reinvestigated by inseminating Strongylocentrotus purpuratus eggs in HCO^?₃-free seawater, then bubbling the seawater with N₂ to remove volatile acid. Fertilization acid production occurred in HCO^?₃-free seawater and with N₂-bubbling, the pH rose 0.28 ± 0.08 unit, significantly less than the rise of 0.63 ± 0.14 unit during N₂-bubbling of HCO^?₃-free seawater that had been acidified with CO₂ and similar to the rise of 0.18 ± 0.07 unit when acidification was with HCl. We conclude that most, if not all, of the sea urchin fertilization acid is nonvolatile and thus is not CO₂; since it is not a weak acid, it must be H⁺.     

Hydrogel design for supporting neurite outgrowth and promoting gene delivery to maximize neurite extension

Hydrogels capable of gene delivery provide a combinatorial approach for nerve regeneration, with the hydrogel supporting neurite outgrowth and gene delivery inducing the expression of inductive factors. This report investigates the design of hydrogels that balance the requirements for supporting neurite growth with those requirements for promoting gene delivery. Enzymatically-degradable PEG hydrogels encapsulating dorsal root ganglia explants, fibroblasts, and lipoplexes encoding nerve growth factor were gelled within channels that can physically guide neurite outgrowth. Transfection of fibroblasts increased with increasing concentration of Arg-Gly-Asp (RGD) cell adhesion sites and decreasing PEG content. The neurite length increased with increasing RGD concentration within 10% PEG hydrogels, yet was maximal within 7.5% PEG hydrogels at intermediate RGD levels. Delivering lipoplexes within the gel produced longer neurites than culture in NGF-supplemented media or co-culture with cells exposed to DNA prior to encapsulation. Hydrogels designed to support neurite outgrowth and deliver gene therapy vectors locally may ultimately be employed to address multiple barriers that limit regeneration.     

Echinostome infection in green frogs (Rana clamitans) is stage and age dependent

The increasing threat of emerging infectious diseases in both wildlife and humans has spurred interest in the causes of disease emergence, including the role of anthropogenic change. A prior field study of infection patterns in amphibians suggests that echinostome infection may be an emerging disease of green frogs Rana clamitans living in urbanized environments. We examined the impact of echinostome infection on green frog tadpoles at a wide range of developmental stages (Gosner stage 25–39). Echinostome infection was associated with green frog mortality rates of up to 40% in an early developmental stage, and none in later developmental stages. Tadpoles exposed to higher echinostome doses exhibited higher edema rates, a potential sign of compromised renal function. Histopathological analysis further supported the hypothesis that echinostome-induced tadpole mortality resulted from compromised renal function. Given that the timing of highest cercarial shedding can coincide with the most vulnerable stages of green frog tadpole development, echinostomes could significantly impact green frog survival in nature.     

Extensive population decline in the Tasmanian devil predates European settlement and devil facial tumour disease

The Tasmanian devil (Sarcophilus harrisii) was widespread in Australia during the Late Pleistocene but is now endemic to the island of Tasmania. Low genetic diversity combined with the spread of devil facial tumour disease have raised concerns for the species’ long-term survival. Here, we investigate the origin of low genetic diversity by inferring the species'' demographic history using temporal sampling with summary statistics, full-likelihood and approximate Bayesian computation methods. Our results show extensive population declines across Tasmania correlating with environmental changes around the last glacial maximum and following unstable climate related to increased ‘El Niño–Southern Oscillation’ activity.