Enzymatic Hydrolysis of Water Lentil (Duckweed): An Emerging Source of Proteins for the Production of Antihypertensive Fractions.

Water lentil (Duckweed), an emerging protein source, is a small floating aquatic plant with agronomic and compositional characteristics rendering it a potential source of bioactive peptides. However, enzymatic hydrolysis of duckweeds has only been carried out to assess the antioxidant and antimicrobial activities of the hydrolysates. The main objectives of this study were to perform enzymatic hydrolysis of duckweed powder utilizing several enzymes and to evaluate the final antihypertensive activity of the fractions. Duckweed powder was efficiently hydrolyzed by pepsin, chymotrypsin, papain and trypsin, with degree of hydrolysis ranging from 3% to 9%, even without prior extraction and concentration of proteins. A total of 485 peptide sequences were identified in the hydrolysates and only 51 were common to two or three hydrolysates. It appeared that phenolic compounds were released through enzymatic hydrolyses and primarily found in the supernatants after centrifugation at concentrations up to 11 mg gallic acid/g sample. The chymotryptic final hydrolysate, the chymotryptic supernatant and the papain supernatant increased the ACE inhibitory activity by more than 6- to 8-folds, resulting in IC50 values ranging between 0.55 to 0.70 mg peptides/mL. Depending on the fraction, the ACE-inhibition was attributed to either bioactive peptides, phenolic compounds or a synergistic effect of both. To the best of our knowledge, this was the first study to investigate the enzymatic hydrolysis of duckweed proteins to produce bioactive peptides with therapeutic applications in mind.

Nisin Purification from a Cell-Free Supernatant by Electrodialysis in a Circular Economy Framework.

Nisin, an antimicrobial peptide produced by Lactococcus lactis strains, is a promising natural preservative for the food industry and an alternative to antibiotics for the pharmaceutical industry against Gram-positive bacteria. Nisin purification is commonly performed using salting out and chromatographic techniques, which are characterized by their low yields, the use of solvents and the production of large volumes of effluents. In the present work, the purification of nisin from a cell-free supernatant (CFS), after the production of nisin by fermentation on a whey permeate medium, was studied using ammonium sulfate precipitation and electrodialysis (ED) as a promising eco-friendly process for nisin purification. Results showed an increase in nisin precipitation using a 40% ammonium sulfate saturation (ASS) level with a purification fold of 73.8 compared with 34.5 and no purification fold for a 60% and 20% ASS level, respectively. The results regarding nisin purification using ED showed an increase in nisin purification and concentration fold, respectively, of 21.8 and 156 when comparing the final product to the initial CFS. Nisin-specific activity increased from 75.9 ± 4.4 to 1652.7 ± 236.8 AU/mg of protein. These results demonstrated the effectiveness of ED coupled with salting out for nisin purification compared with common techniques. Furthermore, the process was noteworthy for its relevance in a circular economy scheme, as it does not require any solvents and avoids generating polluting effluents. It can be employed for the purification of nisin and the recovery of salts from salting out, facilitating their reuse in a circular economy.

How Discoloration of Porcine Cruor Hydrolysate Allowed the Identification of New Antifungal Peptides.

Porcine blood is an important by-product from slaughterhouses and an abundant source of proteins. Indeed, cruor, the solid part of blood, is mainly composed of hemoglobin. Its enzymatic hydrolysis with pepsin generates a diversity of peptides, particularly antimicrobials. One of the downsides of using these hydrolysates as food bio-preservatives is the color brought by the heme, which can be removed by discoloration. Nonetheless, the effects of this procedure on the antimicrobial peptide population have not been completely investigated. In this study, its impacts were evaluated on the final antibacterial and antifungal activities of a cruor hydrolysate. The results demonstrated that 38 identified and characterized peptides showed a partial or total decrease in the hydrolysate, after discoloration. Antifungal activities were observed for the raw and discolored hydrolysates: MICs vary between 0.1 and 30.0 mg/mL of proteins, and significant differences were detected between both hydrolysates for the strains S. boulardii, C. guilliermondii, K. marxianus, M. racemosus and P. chrysogenum. The raw hydrolysate showed up to 12 times higher antifungal activities. Hence, peptides with the highest relative abundance decrease after discoloration were synthesized and tested individually. In total, eight new antifungal peptides were characterized as active and promising. To our knowledge, this is the first time that effective antifungal peptide sequences have been reported from porcine cruor hydrolysates.

Impact of Pulsed Electric Fields and pH on Enzyme Inactivation and Bioactivities of Peptic Hydrolysates Produced from Bovine and Porcine Hemoglobin.

The production of bioactive peptides from hemoglobin via peptic hydrolysis is a promising alternative to valorizing slaughterhouse blood proteins. Nevertheless, it has some limitations such as low yield, high cost of enzymes, and the use of chemical reagents. The latter is aggravated by the pH increase to inactivate the enzyme, which can affect the bioactivity of the peptides. Thus, this study aimed to evaluate the effect of pulsed electric fields (PEF) on the pepsin inactivation and biological activities (antimicrobial and antioxidant) of hemoglobin hydrolysates. Bovine (Hb-B) and porcine (Hb-P) hemoglobin were hydrolyzed with pepsin for 3 h and treated with PEFs to inactivate the enzyme. The degree of hydrolysis (DH) did not show significant changes after PEF inactivation, whereas peptide population analysis showed some changes in PEF-treated hydrolysates over time, suggesting residual pepsin activity. PEF treatments showed no significant positive or negative impact on antimicrobial and antioxidant activities. Additionally, the impact of pH (3, 7, and 10) on bioactivity was studied. Higher pH fostered stronger anti-yeast activity and DPPH-scavenging capacity, whereas pH 7 fostered antifungal activity. Thus, the use of hemoglobin from the meat industry combined with PEF treatments could fit the circular economy concept since bioactive peptides can be produced more eco-efficiently and recycled to reduce the spoilage of meat products. Nevertheless, further studies on PEF conditions must be carried out to achieve complete inactivation of pepsin and the potential enhancement of peptides' bioactivity.

Identification of Gastrointestinal Digestion Stable Antihypertensive Fish Peptides from Atlantic Mackerel (Scomber scombrus).

High blood pressure has been recognized as one of the main risk factors of cardiometabolic syndrome by health organizations. Hypertension is medicated by various classes of synthetic drugs; however, adverse effects have repeatedly been reported. Moreover, natural alternatives such as fish peptides have been effective in the treatment and prevention of hypertension. The aim of our study was to fractionate and identify antihypertensive peptides. Fractions were produced using different techniques of solid-phase extraction (SPE), pressure-driven ultrafiltration (UF), and electro04dialysis with UF membrane. According to our results, the hydrophobic fraction of SPE (IC50 5 µg) was the most potent anti-angiotensin converting enzyme (ACE) product. Findings of the study suggest that the separation technique plays an important role in the isolation efficiency of antihypertensive biopeptides. Importantly, the hydrophobic fraction's activity was retained through a static model of an in vitro gastrointestinal digestion system. In conclusion, polarity regardless of charge and size was the most important factor for anti-ACE activity of an Atlantic mackerel biopeptide. In addition, the presence of leucine at either of the extremities (C- or N-terminal) and/or leucine-rich motifs could well explain the hypotensive effect of our active fraction.

Production of Demineralized Antibacterial, Antifungal and Antioxidant Peptides from Bovine Hemoglobin Using an Optimized Multiple-Step System: Electrodialysis with Bipolar Membrane.

Numerous studies have shown that bovine hemoglobin, a protein from slaughterhouse waste, has important biological potential after conventional enzymatic hydrolysis. However, the active peptides could not be considered pure since they contained mineral salts. Therefore, an optimized multi-step process of electrodialysis with bipolar membranes (EDBM) was carried out to produce discolored and demineralized peptides without the addition of chemical agents. The aim of this study was to test the antibacterial, antifungal and antioxidant activities of discolored and demineralized bovine hemoglobin hydrolysates recovered by EDBM and to compare them with raw and discolored hydrolysates derived from conventional hydrolysis. The results demonstrate that discolored-demineralized hydrolysates recovered from EDBM had significant antimicrobial activity against many bacterial (gram-positive and gram-negative) and fungal (molds and yeast) strains. Concerning antibacterial activity, lower MIC values for hydrolysates were registered against Staphylococcus aureus, Kocuria rhizophila and Listeria monocytogenes. For antifungal activity, lower MIC values for hydrolysates were registered against Paecilomyces spp., Rhodotorula mucilaginosa and Mucor racemosus. Hemoglobin hydrolysates showed fungicidal mechanisms towards these fungal strains since the MFC/MIC ratio was ≤4. The hydrolysates also showed a potent antioxidant effect in four different antioxidant tests. Consequently, they can be considered promising natural, low-salt food preservatives. To the best of our knowledge, no previous studies have identified the biological properties of discolored and demineralized bovine hemoglobin hydrolysates.

Semi-Industrial Production of a DPP-IV and ACE Inhibitory Peptide Fraction from Whey Protein Concentrate Hydrolysate by Electrodialysis with Ultrafiltration Membrane.

The separation by electrodialysis with ultrafiltration membranes (EDUF), at a semi-industrial scale, of a new whey protein hydrolysate obtained from a whey protein concentrate was assessed. After 6 h of treatment, more than 9 g of peptides were recovered in the peptide recovery fraction, for a recovery yield of 5.46 ± 0.56% and containing 18 major components. Among these components, positively charged peptides, such as ALPMHIR + PHMIR, LIVTQTMK and TKIPAVF, were present, and their relative abundances increased by nearly 1.25 X and up to 7.55 X. The presence of these peptides may be promising, as ALPMHIR has a strong activity against angiotensin-converting enzyme (ACE), and LIVTQTMK has structural properties that could interfere with dipeptidyl peptidase-IV (DPP-IV). Many neutral peptides were also recovered alongside those. Nevertheless, the inhibitory activity against DPP-IV and ACE increased from 2 X and 4 X, respectively, in the peptide recovery fraction compared to the initial hydrolysate, due to the improved content in bioactive peptides. Thus, this new hydrolysate is well-suited for the large-scale production of a peptide fraction with high bioactivities. Furthermore, what was achieved in this work came close to what could be achieved for the industrial production of a bioactive peptide fraction from whey proteins.

Scale-Up and Long-Term Study of Electrodialysis with Ultrafiltration Membrane for the Separation of a Herring Milt Hydrolysate.

Electrodialysis with ultrafiltration membrane (EDUF) was selected to separate a herring milt hydrolysate (HMH) in a scale-up and long-term study for the recovery of bioactive peptides. The scale-up was performed to maximise peptide recovery by placing a total membrane area of 0.08 m2 for each anionic and cationic compartment. Twelve consecutive runs were carried out, for a total of 69 h, with minimal salt solution cleaning in between experiments. The final peptide migration rate showed that cationic peptides had a higher average migration rate (5.2 ± 0.8 g/m2·h), compared to anionic peptides (4.7 ± 1.1 g/m2·h). Migration was also selective according to peptide identifications and molecular mass distribution where only small molecular weights were found (<1000 Da) in both recovery compartments. The areal system resistance slightly decreased during each run and the averaged values were stable in between experiments since they were all found in the 95% confidence interval. In addition, total relative energy consumption was quite consistent with an average value of 39.95 ± 6.47 Wh/g all along the 12 consecutive runs. Finally, according to membrane characterization, there was no visual fouling on the different membranes present in the EDUF cell after 69 h of treatment. This may be due to the salt cleaning in between experiments which allowed removal of peptides from the membranes, thus allowing recovering initial system working parameters at the beginning of each run. The entire process was revealed to be very consistent and repeatable in terms of peptide migration, global system resistance, and energy consumption. To the best of our knowledge, this is the first time such EDUF conditions (membrane surface, duration, and minimal salt cleaning between experiments) are being tested on a complex hydrolysate.

Glucoregulatory and Anti-Inflammatory Activities of Peptide Fractions Separated by Electrodialysis with Ultrafiltration Membranes from Salmon Protein Hydrolysate and Identification of Four Novel Glucoregulatory Peptides.

Natural bioactive peptides are suitable candidates for preventing the development of Type 2 diabetes (T2D), by reducing the various risk factors. The aim of this study was to concentrate glucoregulatory and anti-inflammatory peptides, from salmon by-products, by electrodialysis with ultrafiltration membrane (EDUF), and to identify peptides responsible for these bioactivities. Two EDUF configurations (1 and 2) were used to concentrate anionic and cationic peptides, respectively. After EDUF separation, two fractions demonstrated interesting properties: the initial fraction of the EDUF configuration 1 and the final fraction of the EDUF configuration 2 both showed biological activities to (1) increase glucose uptake in L6 muscle cells in insulin condition at 1 ng/mL (by 12% and 21%, respectively), (2) decrease hepatic glucose production in hepatic cells at 1 ng/mL in basal (17% and 16%, respectively), and insulin (25% and 34%, respectively) conditions, and (3) decrease LPS-induced inflammation in macrophages at 1 g/mL (45% and 30%, respectively). More impressive, the initial fraction of the EDUF configuration 1 (45% reduction) showed the same effect as the phenformin at 10 µM (40%), a drug used to treat T2D. Thirteen peptides were identified, chemically synthesized, and tested in-vitro for these three bioactivities. Thus, four new bioactive peptides were identified: IPVE increased glucose uptake by muscle cells, IVDI and IEGTL decreased hepatic glucose production (HGP) of insulin, whereas VAPEEHPTL decreased HGP under both basal condition and in the presence of insulin. To the best of our knowledge, this is the first time that (1) bioactive peptide fractions generated after separation by EDUF were demonstrated to be bioactive on three different criteria; all involved in the T2D, and (2) potential sequences involved in the improvement of glucose uptake and/or in the regulation of HGP were identified from a salmon protein hydrolysate.

Bovine Hemoglobin Enzymatic Hydrolysis by a New Eco-Efficient Process-Part II: Production of Bioactive Peptides.

Bovine cruor, a slaughterhouse waste, was mainly composed of hemoglobin, a protein rich in antibacterial and antioxidant peptides after its hydrolysis. In the current context of food safety, such bioactive peptides derived from enzymatic hydrolysis of hemoglobin represent potential promising preservatives for the food sector. In this work, the hemoglobin hydrolysis to produce bioactive peptides was performed in a regulated pH medium without the use of chemical solvents and by an eco-efficient process: electrodialysis with bipolar membrane (EDBM). Bipolar/monopolar (anionic or cationic) configuration using the H+ and OH- generated by the bipolar membranes to regulate the pH was investigated. The aim of this study was to present and identify the bioactive peptides produced by EDBM in comparison with conventional hydrolysis and to identify their biological activity. The use of the EDBM for the enzymatic hydrolysis of hemoglobin has allowed for the production and identification of 17 bioactive peptides. Hydrolysates obtained by EDBM showed an excellent antimicrobial activity against six strains, antioxidant activity measured by four different tests and for the first time anti-fungal activities against five yeasts and mold strains. Consequently, this enzymatic hydrolysis carried out in regulated pH medium with bipolar membranes could provide bioactive peptides presenting antibacterial, antifungal and antioxidant interest.

Bovine Hemoglobin Enzymatic Hydrolysis by a New Ecoefficient Process-Part I: Feasibility of Electrodialysis with Bipolar Membrane and Production of Neokyotorphin (α137-141).

Neokyotorphin (α137-141) is recognized as an antimicrobial peptide and a natural meat preservative. It is produced by conventional enzymatic hydrolysis of bovine hemoglobin, a major component of cruor, a by-product of slaughterhouses. However, during conventional hydrolysis, chemical agents are necessary to adjust and regulate the pH of the protein solution and the mineral salt content of the final hydrolysate is consequently high. To produce this peptide of interest without chemical agents and with a low salt concentration, electrodialysis with bipolar membrane (EDBM), an electromembrane process recognized as a green process, with two different membrane configurations (cationic (MCP) and anionic (AEM) membranes) was investigated. Hydrolysis in EDBM showed the same enzymatic mechanism, "Zipper", and allowed the generation of α137-141 in the same concentration as observed in conventional hydrolysis (control). EDBM-MCP allowed the production of hydrolysates containing a low concentration of mineral salts but with fouling formation on MCP, while EDBM-AEM allowed the production of hydrolysates without fouling but with a similar salt concentration than the control. To the best of our knowledge, this was the first time that EDBM was demonstrated as a feasible and innovative technology to produce peptide hydrolysates from enzymatic hydrolysis.

How Molecular Weight Cut-Offs and Physicochemical Properties of Polyether Sulfone Membranes Affect Peptide Migration and Selectivity during Electrodialysis with Filtration Membranes.

Filtration membranes (FMs) are an integral part of electrodialysis with filtration membranes (EDFM), a green and promising technology for bioactive peptide fractionation. Therefore, it is paramount to understand how physicochemical properties of FMs impact global and selective peptide migration to anionic (A-RC) and cationic (C+RC) peptide recovery compartments during their simultaneous separation by EDFM. In this context, six polyether sulfone (PES) membranes with molecular weight cut-offs (MWCO) of 5, 10, 20, 50, 100 and 300 kDa were characterized and used during EDFM to separate peptides from a complex whey protein hydrolysate. Surface charge, roughness, thickness and surface/pores nature of studied PES membranes were similar with small differences in conductivity, porosity and pore size distribution. Interestingly, global peptides migration to both recovery compartments increased linearly as a function of MWCO. However, peptide selectivity changed according to the recovery compartments and/or the peptide's charge and MW with an increase in MWCO of FMs. Indeed, in A-RC, the relative abundance (RA) of peptides having low negative charge and MW (IDALNENK and VLVLDTDYK) decreased (45% to 19%) with an increase in MWCO, while the opposite for peptides having high negative charge and MW (TPEVDDEALEK, TPEVDDEALEKFDK & VYVEELKPTPEGDLEILLQK) (increased from 16% to 43%). Concurrently, in C+RC, regardless of MWCO used, the highest RA was observed for peptides having low positive charge and MW (IPAVFK & ALPMHIR). It was the first time that the significant impact of charge, MWCO and pore size distribution of PES membranes on a wide range of MWCO was demonstrated on EDFM performances.

How Charge and Triple Size-Selective Membrane Separation of Peptides from Salmon Protein Hydrolysate Orientate their Biological Response on Glucose Uptake.

The valorization of by-products from natural organic sources is an international priority to respond to environmental and economic challenges. In this context, electrodialysis with filtration membrane (EDFM), a green and ultra-selective process, was used to separate peptides from salmon frame protein hydrolysate. For the first time, the simultaneous separation of peptides by three ultrafiltration membranes of different molecular-weight exclusion limits (50, 20, and 5 kDa) stacked in an electrodialysis system, allowed for the generation of specific cationic and anionic fractions with different molecular weight profiles and bioactivity responses. Significant decreases in peptide recovery, yield, and molecular weight (MW) range were observed in the recovery compartments depending on whether peptides had to cross one, two, or three ultrafiltration membranes. Moreover, the Cationic Recovery Compartment 1 fraction demonstrated the highest increase (42%) in glucose uptake on L6 muscle cells. While, in the anionic configuration, both Anionic Recovery Compartment 2 and Anionic Recovery Compartment 3 fractions presented a glucose uptake response in basal condition similar to the insulin control. Furthermore, Cationic Recovery Compartment 3 was found to contain inhibitory peptides. Finally, LC-MS analyses of the bioassay-guided bioactive fractions allowed us to identify 11 peptides from salmon by-products that are potentially responsible for the glucose uptake improvement.

Evidence of anti-proliferative activities in blue mussel (Mytilus edulis) by-products.

Shellfish waste components contain significant levels of high quality protein and are therefore a potential source for biofunctional high-value peptides. The feasibility of applying a pilot scale enzymatic hydrolysis process to whole Mytilus edulis and, by fractionation, recover hydrolysates presenting a biological activity of interest, was evaluated. Fractions were tested on four immortalized cancerous cell lines: A549, BT549, HCT15 and PC3. The 50 kDa fraction, enriched in peptides, presented anti-proliferative activity with all cell lines and results suggest a bioactive molecule synergy within the fraction. At a protein concentration of 44 µg/mL, the 50 kDa fraction induced a mortality of 90% for PC3, 89% for A549, 85% for HCT15 and of 81% for BT549 cell lines. At the low protein concentration of only 11 µg/mL the 50 kDa fraction still entails a cell mortality of 76% for A549 and 87% for PC3 cell lines. The 50 kDa fraction contains 56% of proteins, 3% of lipids and 6% of minerals on a dry weight basis and the lowest levels detected of taurine and methionine and highest levels of threonine, proline and glycine amino acids. The enzymatic hydrolysis process suggests that Mytilus edulis by-products should be viewed as high-valued products with strong potential as anti-proliferative agent and promising active ingredients in functional foods.

Evidence of Antibacterial Activities in Peptide Fractions Originating from Snow Crab (Chionoecetes opilio) By-Products.

Antibacterial peptide fractions generated via proteolytic processing of snow crab by-products exhibited activity against Gram-negative and Gram-positive bacteria. Among the bacterial strains tested, peptide fractions demonstrated inhibitory activity against the Gram-negative bacteria such as Aeromonas caviae, Aeromonas hydrophila, Campylobacter jejuni, Listonella anguillarum, Morganella morganii, Shewanella putrefasciens, Vibrio parahaemolyticus and Vibrio vulnificus and against a few Gram-positive bacteria such as Listeria monocytogenes, Staphylococcus epidermidis and Streptococcus agalactiae. The principal bioactive peptide fraction was comprised mainly of proteins and minerals (74.3 and 15.5%, respectively). Lipids were not detected. The amino acid content revealed that arginine (4.6%), glutamic acid (5.3%) and tyrosine (4.8%) residues were represented in the highest composition in the antibacterial peptide fraction. The optimal inhibitory activity was observed at alkaline pH. The V. vulnificus strain, most sensitive to the peptide fraction, was used to develop purification methods. The most promising chromatography resins selected for purification, in order to isolate peptides of interest and to carry out their detailed biochemical characterization, were the SP-Sepharose™ Fast Flow cation exchanger and the Phenyl Sepharose™ High Performance hydrophobic interaction media. The partially purified antibacterial peptide fraction was analyzed for minimum inhibitory concentration (MIC) determination, and the value obtained was 25 µg ml(-1). Following mass spectrometry analysis, the active peptide fraction seems to be a complex of molecules comprised of several amino acids and other organic compounds. In addition, copper was the main metal found in the active peptide fraction. Results indicate the production of antibacterial molecules from crustacean by-products that support further applications for high-value bioproducts in several areas such as food and health.

Pelagic fish hydrolysates as peptones for bacterial culture media.

For several years in the Quebec fisheries' industry, landings of pelagic fish have been calculated at over 4000 tons. These under-exploited species, rich in lipids and proteins, could be used in valuable new products. In the present study, hydrolysates of mackerel and herring were produced and utilized as sources of peptones in the formulation of new bacterial culture media. The molecular weight distribution analysis showed that molecules present in the hydrolysates were lower than 1300 Da for herring, and lower than 930 Da for mackerel. The formulated media were compared with reference media using 6 bacterial strains (3 lactic acid (LAB) and 3 non-lactic). The absorbance (OD) and carbohydrate measurements revealed that the formulated media possessed similar yields in comparison with the reference media. Finally, the inhibition of Listeria innocua by LAB bacteriocins was evaluated. Results obtained for Pediococcus acidilactici demonstrated high activities for each medium studied. Thus, the medium containing herring peptones generated the highest bacteriocin titre (32768 AU/mL), followed by both the medium containing mackerel peptones and the MRS7 medium (16384 AU/mL). Each medium containing the fish hydrolysates efficiently supported the growth of the bacterial strains. Pelagic fish peptones are promising as a novel bacterial culture media.

Characterization of enzymatic hydrolyzed snow crab (Chionoecetes opilio) by-product fractions: a source of high-valued biomolecules.

Snow crab (Chionoecetes opilio) constitutes valuable and nutritional sources of components, such as proteins, lipids and chitin. The present investigation was undertaken to evaluate the feasibility of applying a pilot scale enzymatic hydrolysis process of snow crab by-products, followed by fractionation, in order to recover enriched high-valued compounds. The yield of snow crab by-products recovered after manual processing; on a dry weight was 87.4%. The by-products (raw materials) were mainly moist (approximately 78%), and contained 42.9% proteins, 14.8% lipids, 25.7% minerals, 16.2% chitin, all expressed on a dry weight. The fatty acid profile of snow crab by-products and all fractions obtained following processing showed a higher content in mono-unsaturated fatty acids (MUFAs; approximately 50%), followed by polyunsaturated fatty acids (PUFAs; approximately 20%) and saturated fatty acids (SFAs; approximately 15%). The n-3/n-6 ratio was approximately 10 and represents a good index of nutritional value for snow crab oil by-products. Most protein enriched fractions demonstrate a well-balanced amino acid composition, notably the most essential amino acids. These protein fractions are characterized by biomolecules having a relatively low molecular weight (35 kDa and less) range. The enzymatic hydrolysis process developed in this study shows that snow crab by-products should to be viewed as having the potential of being identified as high-valued products. Even though the process could be optimized, it is controllable, and depending on hydrolyses conditions, the products obtained are reproducible and well defined. Results presented in this study indicate that snow crab by-products may serve as excellent nutritional components for future applications in the health and food sectors.

Purification, cloning, and sequencing of a 3,5-dichlorophenol reductive dehalogenase from Desulfitobacterium frappieri PCP-1.

A membrane-associated 3,5-dichlorophenol reductive dehalogenase was isolated from Desulfitobacterium frappieri PCP-1. The highest dehalogenase activity was observed with the biomass cultured at 22 degrees C, compared to 30 and 37 degrees C, where the cell suspensions were 2.2 and 9.6 times less active, respectively. The reductive dehalogenase was purified 12.7-fold to apparent homogeneity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single band with an apparent molecular mass of 57 kDa. Its dechlorinating activity was not inhibited by sulfate and nitrate but was completely inhibited by 2.5 mM sulfite and 10 mM KCN. A mixture of iodopropane and titanium citrate caused a light-reversible inhibition of the dechlorinating activities, suggesting the involvement of a corrinoid cofactor. Several polychlorophenols were dechlorinated at the meta and para positions. The apparent K(m) for 3,5-dicholorophenol was 49.3 +/- 3.1 microM at a methyl viologen concentration of 2 mM. Six internal tryptic peptides were sequenced by mass spectrometry. One open reading frame (ORF) was found in the Desulfitobacterium hafniense genome containing these peptide sequences. This ORF corresponds to a gene coding for a CprA-type reductive dehalogenase. The corresponding ORF (named cprA5) in D. frappieri PCP-1 was cloned and sequenced. The cprA5 gene codes for a 548-amino-acid protein that contains a twin-arginine-type signal for secretion. The gene product has a cobalamin binding site motif and two iron-sulfur binding motifs and shows 66% identity (76 to 77% similarity) with some tetrachloroethene reductive dehalogenases. This is the first CprA-type reductive dehalogenase that can dechlorinate chlorophenols at the meta and para positions.

Continuous renal replacement therapy after cardiac surgery. Review of 85 cases.

BACKGROUND/AIMS: To evaluate the outcome of patients who require continuous renal replacement therapy (CRRT) following cardiac surgery. METHODS: All patients who received CRRT after cardiac surgery over more than 4 years at the Surgical Intensive Care Unit of the Montreal Heart Institute were reviewed. Among 5,564 consecutive patients, 85 underwent CRRT postoperatively. RESULTS: The mean delay between surgery and CRRT initiation was 5 days, and the duration of CRRT was 9 days, without a difference between survivors and non-survivors. Delivered clearances with CRRT were estimated at 25-28 ml/min (approximately 40 liters/day), 29-32 ml/min (approximately 46 liters/day) and 17 ml/min (approximately 25 liters/day) for continuous veno-venous hemofiltration, continuous veno-venous hemodiafiltration and continuous veno-venous hemodialysis, respectively. In-hospital mortality was 43.5%. No difference in mortality was observed between patients with normal renal function at baseline and those with pre-operative renal dysfunction. Mortality was 33.3% after a coronary artery bypass graft (CABG), 57.1% after CABG and valve surgery, 60% after valve surgery, and 72.7% for redo-CABG or redo-valve surgery. 79% of survivors and 86% of non-survivors had received a cardiopulmonary bypass (p = NS). The Simplified Acute Physiology Score II upon intensive care unit (ICU) admission and the requirement of an intra-aortic balloon pump were higher in non-survivors (p < 0.05). The mean length of ICU and hospital stay was 27.4 and 34.2 days for survivors and 17.9 and 22.3 days for non-survivors, respectively (p < 0.05). CONCLUSIONS: Renal impairment is relatively common after cardiac surgery. The mortality of patients who required CRRT after cardiac surgery was 43.5% and was particularly influenced by the type of surgery.