Effect of tandem repeats of antimicrobial peptide CC34 on production of target proteins and activity of Pichia pastoris.

Antimicrobial peptides (AMPs) are attracting attention in the fields of medicine, food, and agriculture because of their broad-spectrum antibacterial properties, low resistance, and low-residue in the body. However, the low yield and instability of the prepared AMP drugs limit their application. In this study, we designed a tetramer of the AMP CC34, constructed and transfected two recombinant expression vectors with pGAPZαA containing a haploid CC34 and tetraploid CC34 (CC34-4js) into Pichia pastoris to explore the effect of biosynthesized peptides. The results showed that CC34 and CC34-4js expression levels were 648.2 and 1105.3 mg/L, respectively, in the fermentation supernatant of P. pastoris. The CC34-4js tetramer showed no antibacterial activity, could be cleaved to the monomer using formic acid, and the hemolytic rate of the polyploid was slightly lower than that of monomeric CC34. The average daily gain, average daily feed intake, feed conversion ratio and immune organ index of rats fed CC34 and CC34-4js showed no differences. In conclusion, CC34-4js exhibited a higher yield and lower hemolysis in P. pastoris than those of CC34. Finally, CC34 and CC34-4js enterokinase lysates showed similar antibacterial activity and both expressed peptides potentially improved the growth performance and organ indices of rats.

[Sedimentary Diatom Records Reveal the Succession of Ecosystem in Lake Xihu, Dali over the Past 50 Years].

In recent decades, intense human activities have caused a decline in many lake ecosystems in Yunnan Province, rendering the transformation of the lake from a clear macrophyte-dominated state to a turbid phytoplankton-dominated state. Improved understanding of the ecological changes in lake ecosystem has significant implications for management. In this study, a small lake in Dali Prefecture of Yunnan Province, i.e., Lake Xihu, was selected. Combined with diatom records and physicochemical proxies from the lake sediments, this paper focuses on the long-term ecological changes in Lake Xihu, Dali since the mid-1960s. The results show that the Lake Xihu, Dali has undergone a significant shift in stable states over the past 50 years. Prior to 2000, the benthic-epiphytic species (i.e., Cocconeis placentula, Staurosira construens, Gomphonema angustum, and Achnanthidium minutissimum) dominated in diatom assemblages, indicating oligotrophic conditions; since 2000, benthic-epiphytic species (i.e., Encyonopsis microcephala and Navicula cryptocephala) and planktonic species (i.e., Cyclotella atomus, Cyclotella meneghiniana, Stephanodiscus hantzschii, and Aulacoseira granulata) dominated successively, indicating mesotrophic to eutrophic conditions. Principal component analysis based on the diatom assemblages in temporal scale showed the response of diatoms succession to nutrients. Redundancy analysis also confirmed that nutrient enrichment was the main driving force for the succession of diatom assemblages in Lake Xihu, Dali. In the past 50 years, climate change and human activities (i.e., agricultural reclamation, fertilization, animal husbandry, and fishery) have enhanced the accumulation of nutrients in the lake. The continuous loading of nutrients promoted the propagation of planktonic algae, and also the productivity of the lake, rendering the transformation to a turbid phytoplankton-dominated state.

An LC/MS quantitative and microdialysis method for cyclovirobuxine D pharmacokinetics in rat plasma and brain: The pharmacokinetic comparison of three different drug delivery routes.

To explore the brain-targeting of cyclovirobuxine D(CVB-D) after administered intranasally, the pharmacokinetics of CVB-D via three different drug delivery routes: intragastric (i.g.), intranasal (i.n.), and intravenous (i.v.) in rat brain and blood was compared. Firstly, an in vivo microdialysis method for sampling CVB-D in both plasma and brain of the rat was established. Secondly, a liquid chromatography-tandem mass spectrometry method has been developed and validated for determination of CVB-D in microdialysis samples. For plasma and brain microdialysis samples, liquid-liquid extraction was used and donepezil was chosen as internal standard. Both were followed by HPLC separation and positive electrospray ionization tandem mass spectrometry detection (ESI-MS/MS). Chromatographic separation was achieved on a agilent C18 column with a mobile phase of methanol-water (50:50, v/v) (pH 3.2) containing 0.1% formic acid and 5mM ammonium acetate. Mass spectrometric detection in the positive ion mode was carried out by selected reaction monitoring (MRM) of the transitions at m/z 403.4→372.3 for CVB-D and m/z 380.2→243.1 for donepezil (IS). Good linearities were obtained in the range of 10-4000ng/mL in rat microdialysates for CVB-D. The lowest limit of quantitation was 5ng/mL, with an extraction recovery >75%, and no significant matrix effects. Intra- and inter-day precisions were all <15% with accuracies of 97.26-116.20%. All of which proved that the established method was successfully applied to the pharmacokinetic study of CVB-D. Simultaneously, brain uptake and pharmacokinetic studies were performed by determination of CVB-D concentration in blood and brain respectively for CVB-D i.g., i.n. and i.v.. Results showed that the intranasal CVB-D could improve brain targeting and had advantages for direct nose to brain transport of CVB-D when compared with injection and oral delivery routes, which indicates that intranasal administration of CVB-D could be a promising approach for the treatment of cerebrovascular disease.