The cloud point was determined for aqueous solutions of four kinds of poly(N-isopropylacrylamide) samples synthesized by radical polymerization in methanol, tert-butanol, benzene, and 1,4-dioxane by the use of azobis(isobutyronitrile) as an initiator, in the range of the weight fraction of the sample from 0.5 to 10%. It has then been found that the cloud points so determined for the samples synthesized in benzene and 1,4-dioxane are definitely lower than those for the samples synthesized in methanol and tert-butanol, although all the samples have almost the same stereochemical composition and the same end group. The observed deviation may be regarded as arising from the difference in the primary structure between the samples.

Most animals show rest/activity rhythms that are regulated by an endogenous timing mechanism, the so-called circadian system. The rhythm becomes weaker with age, but the mechanism underlying the age-associated rhythm change remains to be elucidated. Here we employed Drosophila melanogaster as a model organism to study the aging effects on the rhythm. We first investigated activity rhythms under light-dark (LD) cycles and constant darkness (DD) in young (1-day-old) and middle-aged (30-, 40-, and 50-day-old) wild-type male flies. The middle-aged flies showed a reduced activity level in comparison with young flies. Additionally, the free-running period significantly lengthened in DD, and the rhythm strength was diminished. Immunohistochemistry against pigment-dispersing factor (PDF), a principal neurotransmitter of the Drosophila clock, revealed that PDF levels declined with age. We also found an attenuation of TIMELESS (TIM) oscillation in the cerebral clock neurons in elder flies. Intriguingly, overexpression of PDF suppressed age-associated changes not only in the period and strength of free-running locomotor rhythms but also in the amplitude of TIM oscillations in many pacemaker neurons in the elder flies, suggesting that the age-dependent PDF decline is responsible for the rhythm attenuation. These results suggest that the age-associated reduction of PDF may cause attenuation of intercellular communication in the circadian neuronal network and of TIM cycling, which may result in the age-related rhythm decay.

Prolylhydroxyproline (Pro-Hyp), which is derived from collagen hydrolysate, has been shown to be beneficial for skin and joint health. However, little is known about the distribution of Pro-Hyp in these tissues. In the present study, we investigated the biodistribution of orally administered [ 14 C]Pro-Hyp in rats. Whole-body autoradiography at 30 min after administration of [ 14 C]Pro-Hyp showed that radioactivity is widely distributed in tissues including skin and articular cartilage, with the highest level of radioactivity observed in the gastric and intestinal walls. Incorporation of radioactivity into cells known to respond to ProHyp such as dermal fibroblasts, synovial cells, chondrocytes, osteoblasts, and osteoclasts was observed. The chemical form of [ 14 C]Pro-Hyp-derived radioactivity detected in the tissues was investigated by thin layer chromatography. The radioactive constituents in cartilage extract were two proline-modified peptides (56%), intact Pro-Hyp (5%), and two nonpeptide metabolites (28%). Similar results were obtained for skin and bone marrow. Plasma analysis at 3 to 30 min post-dose suggested that the majority of Pro-Hyp is modified in its proline residue by a first-pass effect without peptide bond hydrolysis. In conclusion, we demonstrated that Pro-Hyp is partly distributed in observed tissues including skin and cartilage in its intact form, which might be responsible for its biological functions.

Transmittance of light passing through an aqueous solution of a linear poly(N-isopropylacrylamide) (PNIPA) sample synthesized by living anionic polymerization was examined in detail in the vicinity of its cloud point. It is found that the transmittance approaches a constant value between 0 and 100% even at a temperature slightly higher than the cloud point, indicating that macroscopic phase separation does not take place at the cloud point in the solution and therefore the cloud-point curve dose not correspond to the binodal. Static and dynamic light scattering measurements were then carried out for aqueous solutions of the linear sample and also of another one synthesized by radical polymerization, which has random-branched structure, at some temperatures considerably lower than the cloud point. It was found that both the PNIPA samples in aqueous solutions form aggregates even at such temperatures, and the number, size, and density profile of the aggregates depend on the kind of chain end group and also on the primary structure.