Physicochemical and functional properties of crawfish chitosan as affected by different processing protocols

Chitosan is made from chitin by a chemical process involving demineralization (DM), deproteinization (DP), decolorization (DC), and deacetylation (DA). Very little work has been done to demonstrate the effects of altering or excluding any of the processing steps on chitosan characteristics. The present study was undertaken to evaluate the effects of process modification during chitosan production on the physiochemical and functional properties of crawfish chitosans. Five experimental chitosan samples (DCMPA, DMCPA, DMPCA, DMPAC, DAMPC) prepared with modified processing protocols and the control (DPMCA - traditional chitosan production process) were evaluated and compared with the two commercial crab chitosans. All samples were subjected to physicochemical (moisture, nitrogen, and ash contents, degree of deacetylation, molecular weight, viscosity, solubility, bulk density, and color) and functional (water binding capacity, fat binding capacity, emulsion capacity, and emulsion viscosity) characteristic analysis. Three experimental replicates were performed with a duplicate analysis of each sample. Results indicated that process modification of crawfish chitosan production yielded some differences on each characteristic compared with the control and commercial chitosans. For instance, changing the sequence of DC for the production of crawfish chitosan affected its properties. DCMPA and DMCPA resulted in an increase in molecular weight and ash, respectively. In contrast, DMPCA led to lower viscosity. The most notable change observed with the DMPAC chitosan was a light brown degraded colored chitosan that exhibited properties of a weak polyelectrolyte. When chitosan process started with DA, a very poor of yield were obtained. When DM and DP were reversed during production, the results showed some difference, e.g., the lower viscosity, higher fat binding capacity, and higher emulsion viscosity of DMPCA over DPMCA. This study demonstrated that process modification of crawfish production affected physicochemical and functional properties. The optimal chitosan production may vary depending on the intended final usages in food systems as demonstrated by functional properties from this study

Diversity of Functional Genes in the Aquatic Nitrogen Cycle

In this PhD thesis the diversity of functional bacterial genes in the nitrogen cycle was investigated with molecular methods in the lakes Plußsee and Schöhsee, and the Baltic Sea. The diversity of ammonia-oxidizing bacteria (AOB) was studied by diversity of specific 16S rDNA and amoA genes. Dominant sequences from Baltic Sea water column and Schöhsee 1 m were related to different Nitrosospira clusters. Sequences from Plußsee 7 m, Schöhsee 12 m and sediment of both lakes were clustered into a purely environmental cluster with no cultivated representatives. Two groups of amoA sequences from Baltic Sea sediment were related to environmental clusters from brackish and marine habitats. The diversity of the evolutionarily related genes for ammonia monooxygenase (AMO) and particulate methane monooxygenase (pMMO) was analyzed. A higher frequency of pmoA sequences, mainly belonging to methane oxidizing bacteria of the gamma subgroup of proteobacteria (g-MOB), was detected. Dominant amoA sequences were related to ammonia oxidizing bacteria of the beta subgroup of proteobacteria (b-AOB), no sequences related to amoA of the γ-AOB were detected. The deduced amino acid sequences of some clones from lake sediments were distantly related to PmoA from Crenothrix polyspora, a filamentous methane oxidizer with an unusual methane monooxygenase. The distribution of denitrifying bacteria was studied by the nitrite reductase genes nirK and nirS. The dominant sequences of nirK from all clone libraries belonged to two distinct phylogenetic clusters, while nirS sequences from both lakes were scattered over several clusters throughout the complete phylogenetic tree, and only few sequences from Baltic Sea overlapped. In the Baltic Sea, nirK-denitrifiers were diverse throughout the water column, while nirS-denitrifiers were dominant in the sediment and almost absent in the water column. In Plußsee the community composition was inverted: nirK-denitrifiers were more diverse in the water column and nirS-denitrifiers in the sediment. In Schöhsee nirS-denitrifiers were highly diverse in water and sediment samples. For nirK and nirS the sequences of the protein were less conserved than those of the gene while the amoA and pmoA protein were conserved, which might be an indication of a differential selection pressure

Distribution of denitrifying bacterial communities in the stratified water column and sediment-water interface in two freshwater lakes and the Baltic Sea

We have studied the distribution and community composition of denitrifying bacteria in the stratified water column and at the sediment-water interface in lakes Plußsee and Schöhsee, and a near-shore site in the Baltic Sea in Germany. Although environmental changes induced by the stratification of the water column in marine environments are known to affect specific populations of denitrifying bacteria, little information is available for stratified freshwater lakes and brackish water. The aim of the present study was to fill this gap and to demonstrate specific distribution patterns of denitrifying bacteria in specific aquatic habitats using two functional markers for the nitrite reductase (nirK and nirS genes) as a proxy for the communities. The leading question to be answered was whether communities containing the genes nirK and nirS have similar, identical, or different distribution patterns, and occupy the same or different ecological niches. The genes nirK and nirS were analyzed by PCR amplification with specific primers followed by terminal restriction fragment length polymorphism (T-RFLP) and by cloning and sequence analysis. Overall, nirS-denitrifiers were more diverse than nirK-denitrifiers. Denitrifying communities in sediments were clearly different from those in the water column in all aquatic systems, regardless of the gene analyzed. A differential distribution of denitrifying assemblages was observed for each particular site. In the Baltic Sea and Lake Plußsee, nirK-denitrifiers were more diverse throughout the water column, while nirS-denitrifiers were more diverse in the sediment. In Lake Schöhsee, nirS-denitrifiers showed high diversity across the whole water body. Habitat-specific clusters of nirS sequences were observed for the freshwater lakes, while nirK sequences from both freshwater lakes and the Baltic Sea were found in common phylogenetic clusters. These results demonstrated differences in the distribution of bacteria containing nirS and those containing nirK indicating that both types of denitrifiers apparently occupy different ecological niche

Comparative analysis of ammonia monooxygenase (amoA) genes in the water column and sediment-water interface of two lakes and the Baltic Sea

The functional gene amoA was used to compare the diversity of ammonia-oxidizing bacteria (AOB) in the water column and sediment-water interface of the two freshwater lakes Plußsee and Schöhsee and the Baltic Sea. Nested amplifications were used to increase the sensitivity of amoA detection, and to amplify a 789-bp fragment from which clone libraries were prepared. The larger part of the sequences was only distantly related to any of the cultured AOB and is considered to represent new clusters of AOB within the Nitrosomonas/Nitrosospira group. Almost all sequences from the water column of the Baltic Sea and from 1-m depth of Schöhsee were related to different Nitrosospira clusters 0 and 2, respectively. The majority of sequences from Plußsee and Schöhsee were associated with sequences from Chesapeake Bay, from a previous study of Plußsee and from rice roots in Nitrosospira-like cluster A, which lacks sequences from Baltic Sea. Two groups of sequences from Baltic Sea sediment were related to clonal sequences from other brackish/marine habitats in the purely environmental Nitrosospira-like cluster B and the Nitrosomonas-like cluster. This confirms previous results from 16S rRNA gene libraries that indicated the existence of hitherto uncultivated AOB in lake and Baltic Sea samples, and showed a differential distribution of AOB along the water column and sediment of these environment

A Synergistic Anti-Diabetic Effect by Ginsenosides Rb1 and Rg3 through Adipogenic and Insulin Signaling Pathways in 3T3-L1 Cells

Although ginsenosides Rb1 and Rg3 have been identified as the significant ginsenosides found in red ginseng that confer anti-diabetic actions, it is unclear whether insulin-sensitizing effects are mediated by the individual compounds or by their combination. To determine the effect of ginsenosides Rb1 and Rg3 on adipocyte differentiation, 3T3-L1 preadipocytes were induced to differentiate the standard hormonal inducers in the absence or presence of ginsenosides Rb1 or Rg3. Additionally, we determined the effects of Rb1, Rg3, or their combination on the expression of genes related to adipocyte differentiation, adipogenic transcription factors, and the insulin signaling pathway in 3T3-L1 cells using semi-quantitative RT-PCR. Rb1 significantly increased the expression of CEBP alpha, PPAR gamma, and aP2 mRNAs. However, Rg3 exerted its maximal stimulatory effect on these genes at 1 mu M concentration, while a high concentration (50 mu M) showed inhibitory effects. Similarly, treatment with Rb1 and Rg3 (1 mu M) increased the expression of IRS-1, Akt, PI3K, GLUT4, and adiponectin. Importantly, co-treatment of Rb1 and Rg3 (9:1) induced the maximal expression levels of these mRNAs. Our data indicate that the anti-diabetic activity of red ginseng is, in part, mediated by synergistic actions of Rb1 and Rg3, further supporting the significance of minor Rg3