[UPLC-Q-TOF-MS-based metabolomics reveals mechanism of Morinda officinalis iridoid glycosides in treating rheumatoid arthritis and bone loss].

This study aimed to investigate the therapeutic effects of Morinda officinalis iridoid glycosides(MOIG) on paw edema and bone loss of rheumatoid arthritis(RA) rats, and analyze its potential mechanism based on ultra-high performance liguid chromatography-guadrupole time-of-flight tandem mass spectrometry(UPLC-Q-TOF-MS) serum metabolomics. RA rats were established by injecting bovin type Ⅱ collagen. The collagen-induced arthritis(CIA) rats were administered drug by gavage for 8 weeks, the arthritic score were used to evaluate the severity of paw edem, serum bone metabolism biochemical parameters were measured by ELISA kits, Masson staining was used to observe the bone microstructure of the femur in CIA rats. UPLC-Q-TOF-MS was used to analyze the alteration of serum metabolite of CIA rats, principal component analysis(PCA) and partial least squares-discriminant analysis(PLS-DA) were used to screen the potential biomarkers, KEGG database analysis were used to construct related metabolic pathways. The results demonstrated that the arthritic score, serum levels of IL-6 and parameters related with bone metabolism including OCN, CTX-Ⅰ, DPD and TRAP were significantly increased, and the ratio of OPG and RANKL was significantly decreased, the microstructure of bone tissue and cartilage were destructed in CIA rats, while MOIG treatments could significantly reduce arthritis score, mitigate the paw edema, reverse the changes of serum biochemical indicators related with bone metabolism, and improve the microstructure of bone tissue and cartilage of CIA rats. The non-targeted metabolomics results showed that 24 altered metabolites were identified in serum of CIA rats; compared with normal group, 13 significantly altered metabolites related to RA were identified in serum of CIA rats, mainly involving alanine, aspartate and glutamate metabolism; compared with CIA model group, MOIG treatment reversed the alteration of 15 differential metabolites, mainly involving into alanine, aspartate and glutamate metabolism, D-glutamine and D-glutamate metabolism, taurine and hypotaurine metabolism, valine, leucine and isoleucine biosynthesis. Therefore, MOIG significantly alleviated paw edema, improved the destruction of microstructure of bone and cartilage in CIA rats maybe through involving into the regulation of amino acid metabolism.

Phylogeography of cultivated and wild ophiopogon japonicus based on chloroplast DNA: exploration of the origin and sustainable cultivation.

BACKGROUND: Ophiopogon japonicus, mainly planted in Sichuan (CMD) and Zhejiang (ZMD) province in China, has a lengthy cultivation history. During the long period of domestication, the genetic diversity of cultivated O. japonicus has substantially declined, which will affect the population continuity and evolutionary potential of this species. Therefore, it is necessary to clarify the phylogeography of cultivated O. japonicus to establish a theoretical basis for the utilization and conservation of the genetic resources of O. japonicus. RESULT: The genetic diversity and population structure of 266 O. japonicus individual plants from 23 sampling sites were analyzed based on 4 chloroplast DNA sequences (atpB-rbcL, rpl16, psbA-trnH and rpl20-5'rps12) to identify the effects of domestication on genetic diversity of cultivars and determine their geographic origins. The results showed that cultivated O. japonicus and wild O. japonicus had 4 and 15 haplotypes respectively. The genetic diversity of two cultivars (Hd = 0.35700, π = 0.06667) was much lower than that of the wild populations (Hd = 0.76200, π = 0.20378), and the level of genetic diversity in CMD (Hd = 0.01900, π = 0.00125) was lower than that in ZMD (Hd = 0.06900, π = 0.01096). There was significant difference in genetic differentiation between the cultivated and the wild (FST = 0.82044), especially between the two cultivars (FST = 0.98254). This species showed a pronounced phylogeographical structure (NST > GST, P < 0.05). The phylogenetic tree showed that the genetic difference between CMD and ZMD was not enough to distinguish the cultivars between the two producing areas by using O. amblyphyllus Wang et Dai as an outgroup. In addition, both CMD and ZMD have a closer relationship with wild populations in Sichuan than that in Zhejiang. The results of the TCS network and species distribution model suggested that the wild population TQ located in Sichuan province could serve as the ancestor of cultivated O. japonicus, which was supported by RASP analysis. CONCLUSION: These results suggest that cultivated O. japonicus has experienced dramatic loss of genetic diversity under anthropogenic influence. The genetic differentiation between CMD and ZMD is likely to be influenced by founder effect and strong artificial selection for plant traits. It appears that wild populations in Sichuan area are involved in the origin of not only CMD but also ZMD. In addition, we also raise some suggestions for planning scientific strategies for resource conservation of O. japonicus based on its genetic diversity and population structure.

[Genetic diversity and genetic structures of original plants of Viticis Fructus:an SSR markers-based analysis].

The present study aimed to provide the protection strategies for wild germplasm resources of original plants of Viticis Fructus and a theoretical basis for the sustainable use of Viticis Fructus. The genetic diversity and genetic structures of the 232 indivi-duals in 19 populations of Vitex rotundifolia and V. trifolia were analyzed by eight SSR markers with tools such as Popgene32, GenAlex 6.502, and STRUCTURE. Bottleneck effect was detected for the population with more than 10 individuals. The results indicated that 42 and 26 alleles were detected from the populations of V. rotundifolia and V. trifolia, respectively, with average expected heterozygo-sities of 0.448 6 and 0.583 9, which are indicative of low genetic diversity. AMOVA revealed the obvious genetic variation of V. rotundifolia and V. trifolia within population(84.43%, P<0.01; 60.37%, P<0.01). Furthermore, in eight SSR loci, six from V. rotundifolia populations and two from V. trifolia populations failed to meet Hardy-Weinberg equilibrium expectations(P<0.05), which confirmed that the populations experienced bottleneck effect. As assessed by Mantel test, geographical distance posed slight impacts on the genetic variation between the populations of V. rotundifolia and V. trifolia. Principal component analysis(PCA) and STRUCTURE analysis demonstrated evident introgression of genes among various populations. The original plants of Viticis Fructus were confirmed low in genetic diversity and genetic differentiation level. Therefore, the protection of wild resources of original plants of Viticis Fructus should be strengthened to ensure its sustainable use.

Isolation, structural properties, bioactivities of polysaccharides from Dendrobium officinale Kimura et. Migo: A review.

Dendrobium officinale Kimura et Migo (D. officinale) is used as herbal medicine and new food resource in China, which is nontoxic and harmless, and can be used as common food. Polysaccharide as one of the main bioactive components in D. officinale, mainly composed of glucose and mannose (Manp: Glcp = 2.01:1.00-8.82:1.00), along with galactose, xylose, arabinose, and rhamnose in different molar ratios and types of glycosidic bonds. Polysaccharides of D. officinale exhibit a variety of biological effects, including immunomodulatory, anti-tumor, gastro-protective, hypoglycemic, anti-inflammatory, hepatoprotective, and vasodilating effects. This paper presents the extraction, purification, structural characteristics, bioactivities, structure-activity relationships and analyzes gaps in the current research on D. officinale polysaccharides. In addition, based on in vitro and in vivo experiments, the possible mechanisms of bioactivities of D. officinale polysaccharides were summarized. We hope that this work may provide helpful references and promising directions for further study and development of D. officinale polysaccharides.

[Partial Nitrification Fast Start-up and Stable Performance of 15℃ SBBR].

In this study, controlled C/N effects on fast start-up and stable performance of partial nitrification process at 15℃ in a Sequenced Batch Biofilm Reactor (SBBR) were investigated. The results showed that partial nitrification successfully fast initiated when C/N was 1.5 but failed when C/N was 0/3 during 60 cycles. Fluorescence in situ hybridization and confocal laser scanning microscope (FISH-CLSM) results showed that ammonia oxidizing bacteria (AOB) was found as the dominant bacteria population when C/N was 1.5. When C/N were 0/3, there were almost no existence of AOB and nitrite oxidative bacteria (NOB). Partial nitrification could be stably achieved without carbon source. However, the addition of an appropriate amount of carbon can effectively improve the nitrification performance, and it is better for the stable operation of partial nitrification. In this experiment, partial nitrification was successfully initiated at high dissolved oxygen (DO) (about 9 mg·L-1) conditions. The average DO was maintained at about 6.5 mg·L-1 during the stable operation, which successfully decoupled partial nitrification from low DO concentration. Excessive residual ammonium concentration in the reactors effectively repressed the growth of NOB and guaranteed the stable operation of partial nitrification. At 15℃, full nitritation was more suitable for sidestream wastewater, while mainstream wastewater was more suitable for partial nitritation.

[Achieving Partial Nitritation in a Continuous-flow Aerobic Granular Sludge Reactor at Different Temperatures Through Ratio Control].

Mature aerobic granular sludge (AGS) was inoculated in a continuous-flow reactor to treat low ammonia sewage, and the feasibility of achieving partial nitritation in a continuous-flow aerobic sludge system and the demand for R value (the ratio of dissolved oxygen and ammonia nitrogen) when partial nitritation is achieved at different temperatures (30, 20, and 10℃) were investigated. The control strategy was designed to maintain a constant ratio between dissolved oxygen (DO) and ammonia nitrogen (NH4+-N). The results revealed that stable nitritation in a continuous-flow aerobic sludge reactor could be achieved via ratio control, and the value of R were 0.50 (±0.05), 0.35 (±0.03), and 0.20 (±0.02) at the temperatures 30, 20, and 10℃, respectively, from which it can be concluded that stronger oxygen-limiting conditions were required when the temperature was lower. The experiment of fluorescence in situ hybridization (FISH) revealed that ammonia oxidizing bacteria (AOB) have a certain concentration, and the relative number of nitrite oxidizing bacteria (NOB) gradually reduced through the ratio control strategy. Based on the ratio control strategy and the characteristics of wastewater quality, full nitritation of high ammonia wastewater may be allowed; however, for low ammonia wastewater, only partial nitritation is recommended.

[Characteristics of Biofilm During the Transition Process of Complete Nitrification and Partial Nitrification].

The objective of the study was to investigate the change of biofilm characteristics when implementing the procedure of partial nitrification. A ratio control strategy (DO/NH4+-N) was taken to achieve partial nitrification, and biofilm samples were obtained at 10.27%, 52.12%, and 93.54% of the nitrite accumulation rate. The amount and spatial distribution of total bacteria, ammonia oxidizing bacteria (AOB), and nitrite oxidative bacteria (NOB) were observed by fluorescence in situ hybridization (FISH) and confocal laser scanning microscope (CLSM) through a three-dimensional excitation emission matrix (EEM) to observe the secretion and composition changes of extracellular polymer substances. Ratio control successfully enriched AOB and achieved partial nitrification under conditions when NOB was not completely washed. Heterotrophic bacteria and nitrifying bacteria coexist in the biofilm. The heterotrophic bacteria were in the outer layer, but nitrifying bacteria were distributed in the biofilm surface at 6-25 µm. During the process of short-range nitrification, the AOB/NOB value gradually increased, and the stable operation period was as high as 15.56. During the operation of the reactor, EPS and microbial flora changes are closely related. When microbial activity decreased, EPS secretion decreased. During the stable operation period of partial nitrification, NOB and other bacteria that are non-resistant to high nitrite nitrous acid declined, and the fluorescence intensity of aromatic protein-like bacteria decreased. However, the three-dimensional fluorescence spectra showed that the chemical composition of EPS was not obvious during the process of partial nitrification.

[Optimization of the Flow Distribution Ratio and Mechanism of Nitrogen Removal in a Multi-level AO Coupled Flow Biochemical Process].

To explore the influence of the influent flow distribution ratio on the denitrification efficiency of low-temperature urban wastewater and analyze the denitrification of multi-level AO coupled flow biochemical process, three-level AO-coupled biofilm technology was used to treat simulated low-C/N urban sewage at a temperature of 10℃±1℃, hydraulic retention time of 8 h, and constant air-water ratio. The reactors were operated under three conditions of inlet water ratios of 5:4:4 (equal volume load), 3:2:1 (equal hydraulic retention time), and 25:15:6 (equal sludge load). The study showed that the multi-level AO-coupled displacement biochemical process has a good removal efficiency with respect to low-temperature and low-C/N wastewater. The pollutant removal efficiency is the highest when the ratio of the influent is 3:2:1 and the average removal rates of COD, NH4+-N, and TN are 87.44%, 96.63%, and 76.81%, respectively. Further studies on the law of nitrogen migration and transformation showed that the main factors constraining the nitrogen removal under three conditions were the nitrification efficiency at each levels, the ratio of 3:2:1 influent reasonably distributing the influent load, and the nitrification efficiency at each level exceeding 85%, creating favorable conditions for denitrification and leading to a higher denitrification efficiency, while the system has the highest total biomass. The research results enrich the theory of multi-level AO cryogenic removal of nitrogen and provide references for engineering designs and applications.

[Comparison of Start-up and Stable Performance of Nitritation in Activated Sludge and Biofilm Processes in a SBR].

This study investigates the effect of increased hydraulic retention time (HRT) and dissolved oxygen (DO) on the destruction and restoration of the performance of activated sludge and biofilm sequencing batch reactor (SBR) nitritation after the processes were simultaneously started up and stably operated at room temperature (25℃). The results showed that was easier to start nitritation in an activated sludge reactor, but nitritation was destroyed when DO was 2-2.5 mg·L-1. The performance could be restored by reducing DO to 0.5-1 mg·L-1. The biofilm process was not affected by DO. Prolonging HRT destroyed the stable operation of the two processes. Compared with the method of biofilm, the activated sludge method had less resistance, but restoring performance was better than for the biofilm process after shortening the HRT. Subsequently, the temperature (20, 15, and 10℃) was continuously reduced to investigate the synergistic effect of DO and temperature on nitritation stabilization. The results showed that the reduction of temperature destroyed the stable operation of nitritation, but the reduction of DO concentration could compensate for the adverse effects of temperature reduction. In addition, when the temperature was higher than 20℃, the rate of nitritation in the activated sludge process was better than that in thebiofilm method. At low temperature (below 15℃), a stable operation of nitritation was easier to achieve with the biofilm method. The capacity of the activated sludge process was hardly changed at 10℃. The above conclusions were confirmed by molecular microbiological analysis, and it was found that the stable operation of nitritation could be achieved without complete wash-out of the nitrite-oxidizing bacteria (NOB).