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Thermal stability is essential for the understanding of protein stability and is a critical quality attribute of therapeutic biologics, including enzymes, fusion proteins, monoclonal antibodies, etc. The commonly used analytical methods, such as differential scanning calorimetry (DSC), differential scanning fluorimetry (DSF), and circular dichroism (CD), have their limitations in measuring protein thermal stability. Through this work, we described a novel method to probe the thermal stability of proteins in various formulations using a temperature scanning viscometer. The viscosity of the material was plotted against the temperature, and the peak in the first derivative of the viscosity versus temperature was shown to be related to the protein melting temperature. The measured melting temperature of bovine serum albumin (BSA) at a concentration of 1 mg/mL in phosphate buffer was 63 °C, which was close to the value of 64 °C obtained by DSC. The unfolding of BSA was confirmed using orthogonal techniques of second derivative ultraviolet-visible (UV-vis) spectroscopy and dynamic light scattering (DLS). This method was also able to reveal the microenvironment changes of proteins, including formulation effects. Other multiple domains proteins including lysozyme and IgG were also tested using this method and showed comparable melting temperatures with DSC. This work showed the feasibility of using a temperature scanning viscometer to measure the thermal stability of proteins in diverse formulation matrices with wider protein concentration ranges.

Species composition and seasonal variations of free-floating Ulva species were investigated in the source area of the world's largest macroalgal blooms during 2009-2015. Based on a combination of a morphological analysis and sequences of nuclear-encoded ITS and 5S rDNA spacer regions, the dominant species in the free-floating Ulva community at the early stage of green tides were Ulva compressa, Ulva flexuosa, and Ulva linza. The first appearance of Ulva prolifera on the sea surface was in mid-May and it dominated the floating Ulva community in June from 2009 to 2011. From 2012-2015, U. prolifera was not only the first species to appear on the sea surface but also the dominant species during the whole early stage of green tides. To explain the successional mechanism, the effects of environmental factors on the growth of four Ulva species were examined in the laboratory under different combinations of light intensity and temperature. It was found that the highest growth rate of U. prolifera was 44.9%/d, which was much higher than the other three Ulva species. The strong tolerance of U. prolifera to extreme conditions also helps it survive and bloom in the Yellow Sea.

Biological products, such as therapeutic proteins, vaccines and cell - based therapeutics have a rapidly growing global market. Monoclonal antibody represents a major portion of the biologics market. For biologics that target gastrointestinal tract, the oral delivery route offers many advantages, such as better patient compliance, easy administration and increased stability, over the parental route of administration. To lay the ground work for the oral delivery of biologics, we studied the solid state properties and effects of compaction pressure, particle size, and storage relative humidity on the stability of immunoglobulin G (IVIG). We employed complementary analytical and biophysical techniques, such as size exclusion chromatography and Dynamic light scattering to characterize the aggregates, circular dichroism and solid state Fourier-transform infrared spectroscopy to evaluate protein secondary structure and nano-DSC to probe thermal stability of protein conformations. Our results showed storage relative humidity could induce conformational changes and aggregation of IVIG. However, the IVIG binding activity did not significantly change with relative humidity. The commonly used compaction pressures did not promote protein aggregation, but noticeably reduced binding activity.

Metabolism, as a ubiquitous biological process, plays a decisive role in the wellbeing of cells, organs, and whole body. Since the endpoint of pharmacological intervention is to either recover or sustain the wellbeing of targeted biological systems, examining metabolic events reveals useful information on diseases and drug treatment. Based on its omics nature and capacity to examine both drug and endogenous metabolism, metabolomics is an ideal systems biology tool to examine chemical and metabolic events in drug discovery and development. Ongoing and potential applications of metabolomics in drug discovery and development include biomarker identification, bioactive compound identification, drug metabolism, metabolic profiling, and mechanistic investigation. Corresponding to four stages of drug discovery and development, why and how metabolomics is used in each stage of drug discovery and development are explained and discussed in four respective sections of this review. Since metabolites, the direct targets of metabolomic analysis, are the end products of gene-, protein-, microbe-, and signal molecule-mediated biological processes, metabolomics possesses many functions that are irreplaceable by other system biology tools. Even though its application is still in the germination phase, metabolomics is a promising platform to bridge chemistry and biology in drug discovery and development.

Identification of multicomponent in traditional Chinese medicine (TCM) is complex and time-consuming. The inspection of the full-scan mass chromatograms was usually performed manually, which is labor-intensive. It is difficult to distinguish low response signals from complex chemical background. Furthermore, this process is typically based on earlier knowledge of the chemical composition of TCM, and those molecules that have not been characterized earlier were thus ignored. In this paper, a strategy using UPLC-MS combined with pattern recognition analysis was developed to simplify and quicken the identification of multicomponent in Abelmoschus manihot (L.) Medik. First, complex signals obtained by UPLC-MS were processed using automated data mining algorithm and further processed with multivariate chemometric methods. Multicomponent in Abelmoschus manihot L. can be clearly displayed in S- and VIP-plot. Using this method, 320 peaks which present in Abelmoschus manihot L. were detected. In the next step, accurate mass spectra of the characteristic markers acquired by QTOF MS were used to estimate their elemental formulae and enable structure identification. By searching in METLIN database, 41 components were tentatively identified in Abelmoschus manihot L. Our results showed that UPLC-MS based-pattern recognition analysis approach can be used to quickly identify TCM multicomponent and for standardization of herbal preparations.

Green algal blooms have occurred in the Yellow Sea for 11 consecutive years since 2007. A “seed bank” comprising micro-propagules including gametes, meiospores, and zygotes, played an important role in the rapid formation of a green tide. In the present study, germination differences among zygotes, meiospores, and gametes were examined. The growth ability and maturation period of alternating generations of sexual Ulva prolifera strains were also assessed. The zygote and meiospore germination rate was 91.67% and 80.29%, respectively, approximately three times greater than that of gametes (30%). In addition, the highest daily growth rate of sporophytes and gametophytes was 266.7% and 288.1%, respectively, and the maturation period of sporophytes and gametophytes was 35.7 and 31.3 days, respectively. These results indicate that sexual reproduction and vegetative growth are mainly responsible for the rapid expansion of macroalgal blooms in the Yellow Sea.

Fecal microbiota transplantation (FMT) has emerged as a highly effective therapy for refractory, recurrent Clostridium difficile infection (CDI), which develops following antibiotic treatments. Intestinal microbiota play a critical role in the metabolism of bile acids in the colon, which in turn have major effects on the lifecycle of C. difficile bacteria. We hypothesized that fecal bile acid composition is altered in patients with recurrent CDI and that FMT results in its normalization. General metabolomics and targeted bile acid analyses were performed on fecal extracts from patients with recurrent CDI treated with FMT and their donors. In addition, 16S rRNA gene sequencing was used to determine the bacterial composition of pre- and post-FMT fecal samples. Taxonomic bacterial composition of fecal samples from FMT recipients showed rapid change and became similar to the donor after the procedure. Pre-FMT fecal samples contained high concentrations of primary bile acids and bile salts, while secondary bile acids were nearly undetectable. In contrast, post-FMT fecal samples contained mostly secondary bile acids, as did non-CDI donor samples. Therefore, our analysis showed that FMT resulted in normalization of fecal bacterial community structure and metabolic composition. Importantly, metabolism of bile salts and primary bile acids to secondary bile acids is disrupted in patients with recurrent CDI, and FMT corrects this abnormality. Since individual bile salts and bile acids have pro-germinant and inhibitory activities, the changes suggest that correction of bile acid metabolism is likely a major mechanism by which FMT results in a cure and prevents recurrence of CDI.

Ethnopharmacological relevance: Abelmoschus manihot (L.) Medik. var. manihot is one of the most commonly used Chinese medicines and has played an important role in treating chronic glomerulonephritis and diabetic nephropathy. Aim of the study: Metabolites identification of traditional Chinese medicine (TCM) is a complex and time-consuming process due to the complicity of TCM and subsequent large number of detected ions. In this paper, UPLC–MS combined with pattern recognition analysis approach were used to simplify and quicken the identification of the metabolites of Abelmoschus Manihot . Materials and methods: Rat urine samples were collected before (as control sample) and after Abelmoschus Manihot administration. Pattern recognition analysis method was used to differentiate components between Abelmoschus Manihot -treated group and its controlled comparison. These components could be considered as Abelmoschus Manihot -related metabolites in vivo . Results: LC–MS based metabolomics could be an advanced tool to help us find metabolites with regards to its capacity of processing large datasets, differentiating and classifying of sample groups, as well as its indiscriminative nature of biomarker and metabolite identification. Using this method, seven metabolites were identified, which are flavonoid aglycone glucuronidation, sulfatation, and methylation metabolites. Conclusion: Our results showed that UPLC–MS based- pattern recognition analysis approach can be used to quickly identify Abelmoschus Manihot related metabolites in biological fluids. Furthermore, this work demonstrates the potential application of combining the UPLC–MS approach with the metabolomics approach in identifying the metabolites of TCM.

In this paper, microdialysis combining with ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) was applied to simultaneously identify components in blood and kidney dialysis after oral administration of Abelmoschus manihot extract. Microdialysis probe was implanted in the jugular vein and the kidney medulla, respectively; microdialysis samples were collected continuously, transferred to microtubes and analyzed by UPLC-Q-TOF/MS. The components in microdialysis samples were separated by an UPLC HSS T3 column and eluted with acetonitrile and water (containing 0.1% formic acid) at a flow rate of 0.4 mL/min. The results showed that unbound constituents in blood circulation of the rat include hyperoside, isoquercitrin, quercetin monoglucuronide, quercetin-3'-O-glucoside, quercetin, myricetin, and hibifolin while unbound constituents in kidney are hyperoside, isoquercitrin, quercetin monoglucuronide, which might be the potential active components in vivo. The developed method was simple and reliable, and could be adopted to rapidly screen and identify potential active components contributing to pharmacological effects of TCM and to better clarify its action mechanism.