Construction of hypoallergenic microgel by soy major allergen ß-conglycinin through enzymatic hydrolysis and lactic acid bacteria fermentation.

Soy allergenicity is a public concern, and the combination of multiple processing methods may be a promising strategy for reducing soy allergenicity. In this study, a novel two-step enzymatic hydrolysis followed by lactic acid bacteria fermentation was proposed for the construction of hypoallergenic soybean protein microgel. ß-Conglycinin was used as the main soy allergen. The effects of different enzymatic hydrolysis (Alcalase, Neutrase, and Protamex) and LAB fermentation on ß-conglycinin microgel formation and its immunoreactivity were investigated. Results showed that the use of different enzymes and the attainment of different degrees of hydrolysis affected the particle distribution and zeta potential in the microgels and leads to differences in microstructure and immunoreactivity. All hydrolysates compared with intact protein accelerated the formation of gel during LAB fermentation. Among the three assayed enzymes, fermented Protamex hydrolysates at 60 min (PF-60) demonstrated a microgel with an overall reduced average particle size (741.20±7.18 nm), lower absolute values of zeta potential (10.43±0.65 mV), and regular gel network. The antigenicity and IgE-binding capacity decreased to the lowest value of 0.30 % and 6.93 %, respectively. Peptidomics and immunoinformatic analysis suggested that PF-60 disrupted 17/30, 16/44, and 23/75 epitopes in the α, α', and ß subunits, respectively. Unlike the LAB-fermented unhydrolyzed ß-conglycinin disrupted epitopes mostly located at the loop domain, PF-60 primarily promoted the exposure and disruption of allergen epitopes with ß-sheet structure located at the core barrel domain. These findings can provide new perspectives on the preparation of hypoallergenic soybean-gel products on edible particulate systems.

Disturbed laterality of non-rapid eye movement sleep oscillations in post-stroke human sleep: a pilot study.

Sleep is known to promote recovery post-stroke. However, there is a paucity of data profiling sleep oscillations in the post-stroke human brain. Recent rodent work showed that resurgence of physiologic spindles coupled to sleep slow oscillations (SOs) and concomitant decrease in pathological delta (δ) waves is associated with sustained motor performance gains during stroke recovery. The goal of this study was to evaluate bilaterality of non-rapid eye movement (NREM) sleep-oscillations (namely SOs, δ-waves, spindles, and their nesting) in post-stroke patients vs. healthy control subjects. We analyzed NREM-marked electroencephalography (EEG) data in hospitalized stroke-patients (n = 5) and healthy subjects (n = 3). We used a laterality index to evaluate symmetry of NREM oscillations across hemispheres. We found that stroke subjects had pronounced asymmetry in the oscillations, with a predominance of SOs, δ-waves, spindles, and nested spindles in affected hemisphere, when compared to the healthy subjects. Recent preclinical work classified SO-nested spindles as restorative post-stroke and δ-wave-nested spindles as pathological. We found that the ratio of SO-nested spindles laterality index to δ-wave-nested spindles laterality index was lower in stroke subjects. Using linear mixed models (which included random effects of concurrent pharmacologic drugs), we found large and medium effect size for δ-wave nested spindle and SO-nested spindle, respectively. Our results in this pilot study indicate that considering laterality index of NREM oscillations might be a useful metric for assessing recovery post-stroke and that factoring in pharmacologic drugs may be important when targeting sleep modulation for neurorehabilitation post-stroke.

Modulation of soy protein immunoreactivity by different matrix structures of lactic acid bacterium-induced soy protein gels: Epitope destruction during in vitro gastroduodenal digestion and absorption.

Soy allergy is a common health problem. Food structure may change the gastroduodenal digestion and absorption of soy proteins, thus leading to the modulation of the immunoreactivity of soy proteins. In this study, lactic acid bacterium (LAB)-fermented soy protein isolates (FSPIs) were prepared at four concentrations (0.2 %-5.0 %, w/v) to present various matrix structures (nongel, NG; weak gel, WG; medium gel, MG; and firm gel, FG) and subjected to in vitro dynamic gastroduodenal digestion model. The results of sandwich enzyme-linked immunosorbent and human serum IgE binding capacity assays demonstrated that FSPI gels, especially the FSPI-MG/WG digestates obtained at the early and medium stages of duodenal digestion (D-5 and D-30), possessed greater potency in immunoreactivity reduction than FSPI-NG and reduced to 1.9 %-68.3 %. The transepithelial transport study revealed that the immunoreactivity of FSPI-MG/WG D-5 and D-30 digestates decreased through the stimulation of interferon-γ production and the induction of dominant Th1/Th2 differentiation. Peptidomics and bioinformatics analyses illustrated that compared with FSPI-NG, the FSPI-gel structure promoted the epitope degradation of the major allergens glycinin G2/G5, ß-conglycinin α/ß subunit, P34, lectin, trypsin inhibitor, and basic 7S globulin. Spatial structure analysis showed that FSPI-gel elicited an overall promotion in the degradation of allergen epitopes located in interior and exterior regions and was dominated by α-helix and ß-sheet secondary structures, whereas FSPI-MG/WG promoted the degradation of epitopes located in the interior region of glycinin/ß-conglycinin and exterior region of P34/basic 7S globulin. This study suggested that the FSPI-gel structure is a promising food matrix for decreasing the allergenic potential of allergenic epitopes during gastroduodenal digestion and provided basic information on the production of hypoallergenic soy products.

Characteristics of gelatin from lizardfish (Saurida micropectoralis) and threadfin bream (Nemipterus hexodon) skins as influenced by extraction conditions.

Gelatins from lizardfish and threadfin bream skins were extracted using distilled water at 45 and 60 °C and 4, 8 and 12 h. Gelatin recovered from both lizardfish and threadfin bream skins was in the range of 63.96-91.46%. As extraction temperature and duration increased, the turbidity of gelatin solution from both species increased. Gelatins isolated from either lizardfish or threadfin bream skins at 45 °C for 4 and 8 h showed the maximum bloom strength (245.03-320.85 g), which were also greater than commercial gelatin from bovine (208.55 g) (P < 0.05). The gelatin gels (6.67%, w/v) could set at 4 °C within 3 min and were able to set at room temperatures within 51.83 min. Gelatins extracted from both fish skins contained α1-, ß- and γ-chains as predominant protein components. The lightness of all gelatin gels faintly declined with an increase in extraction temperature and time. Among the various production conditions explored, lizardfish/threadfin bream skin gelatin developed at 45 °C and 8 h was found to be highly comparable to commercial bovine gelatin. Based on the results obtained, gelatin from both fish species could be used as a replacement for land animal counterparts and can be used in many different food and pharmaceutical products.

Partitioning and recovery of proteases from lizardfish (Saurida micropectoralis) stomach using combined phase partitioning systems and their storage stability.

Partitioning and recovery of proteases from stomach extract (SE) and acidified stomach extract (ASE) of lizardfish using a three-phase partitioning (TPP) system in combination with an aqueous two-phase system (ATPS) was optimized. The highest yield and purity were obtained in the interphase of the TPP system, which consisted of a SE or ASE to t-butanol ratio of 1.0 : 0.5 in the presence of 40% (w/w) (NH4)2SO4. Both TPP fractions were further subjected to ATPS. Phase compositions of ATPS including PEG molecular mass and concentrations as well as types and concentrations of salts influenced protein partitioning. The best ATPS conditions for protease partitioning into the top phase from TPP fractions of SE and ASE were 15% Na3C6H5O7-20% PEG1000 and 20% Na3C6H5O7-15% PEG1000, which increased the purity by 4-fold and 5-fold with the recovered activity of 82% and 77%, respectively. ATPS fractions of SE and ASE were subsequently mixed with several PEGs and salts for back extraction (BE). BE using 25% PEG8000-5% Na3C6H5O7 gave the highest PF and yield for both ATPS fractions. SDS-PAGE investigation revealed that the decrease in contaminating protein bands was observed after the combined partitioning systems. BE fractions of SE and ASE were quite stable at -20 and 0 °C up to 14 days. Therefore, the combination of TPP, ATPS and BE could be effectively applied to recover and purify proteases from the stomach of lizardfish.

Disturbed laterality of non-rapid eye movement sleep oscillations in post-stroke human sleep: a pilot study.

Sleep is known to promote recovery post-stroke. However, there is a paucity of data profiling sleep oscillations post-stroke in the human brain. Recent rodent work showed that resurgence of physiologic spindles coupled to sleep slow oscillations(SOs) and concomitant decrease in pathological delta(δ) waves is associated with sustained motor performance gains during stroke recovery. The goal of this study was to evaluate bilaterality of non-rapid eye movement (NREM) sleep-oscillations (namely SOs, δ-waves, spindles and their nesting) in post-stroke patients versus healthy control subjects. We analyzed NREM-marked electroencephalography (EEG) data in hospitalized stroke-patients (n=5) and healthy subjects (n=3) from an open-sourced dataset. We used a laterality index to evaluate symmetry of NREM oscillations across hemispheres. We found that stroke subjects had pronounced asymmetry in the oscillations, with a predominance of SOs, δ-waves, spindles and nested spindles in one hemisphere, when compared to the healthy subjects. Recent preclinical work classified SO-nested spindles as restorative post-stroke and δ-wave-nested spindles as pathological. We found that the ratio of SO-nested spindles laterality index to δ-wave-nested spindles laterality index was lower in stroke subjects. Using linear mixed models (which included random effects of concurrent pharmacologic drugs), we found large and medium effect size for δ-wave nested spindle and SO-nested spindle, respectively. Our results indicate considering laterality index of NREM oscillations might be a useful metric for assessing recovery post-stroke and that factoring in pharmacologic drugs may be important when targeting sleep modulation for neurorehabilitation post-stroke.

Pectin from plantain peels: Green recovery for transformation into reinforced packaging films.

Plantain peels as agro-waste are generated in the millions of tons per year with no profitable management strategies. On the other hand, the excessive use of plastic packaging threatens the environment and human health. This research aimed to address both problems via a green approach. High-quality pectin was recovered from plantain peels via an enzyme-assisted and ethanol-recycling process. The yield and galacturonic acid (GalA) content of the recovered low methoxy pectin was 12.43% and 25.0%, respectively, when cellulase was added at 50 U per 5 g peel powder, with a significantly higher recovery rate and purity than the pectin products extracted with no cellulase (P ≤ 0.05). The recovered pectin was further integrated and reinforced with beeswax solid-lipid nanoparticles (BSLNs) to fabricate films as a potential alternative packaging material to single-use plastics. The reinforced pectin films showed improved light barrier, water resistance, mechanical, conformational, and morphological properties. This study presents a sustainable strategy to transform plantain peels into pectin products and pectin-based packaging films with broad applications.

Evaluation of the Preservation and Digestion of Seal Meat Processed with Heating and Antioxidant Seal Meat Hydrolysates.

Seal meat is of high nutritive value but is not highly exploited for human food due to ethical issues, undesirable flavors, and loss of nutrients during the processing/cooking step. In this work, commercially available processed seal meat was treated with its hydrolysates as preservatives with the aim of improving nutrient bioavailability. The contents of the nutrients were analyzed after digestion using a simulated dynamic digestion model, and the effects of different processing conditions, i.e., low-temperature processing and storage (25 °C) and high-temperature cooking (100 °C), of seal meat were investigated. Hydrolysates with antioxidant activity decreased the amounts of the less desirable Fe3+ ions in the seal meat digests. After treatment with hydrolysates at room temperature, a much higher total Fe content of 781.99 mg/kg was observed compared to other treatment conditions. The release of amino acids increased with temperature and was 520.54 mg/g for the hydrolysate-treated sample versus 413.12 mg/g for the control seal meat sample treated in buffer. Overall, this study provides useful data on the potential use of seal meat as a food product with high nutritive value and seal meat hydrolysates with antioxidant activity as preservatives to control oxidation in food.

Enzymological characteristics of pepsinogens and pepsins purified from lizardfish (Saurida micropectoralis) stomach.

One major pepsinogen, PG-I, and two minor pepsinogens, PG-II and PG-III were purified from lizardfish stomach by ammonium sulfate precipitation and two chromatographic columns. The three purified PGs migrated as single bands in native-PAGE gels with molecular weights (MW) ranging from 36 to 38 kDa. Each PG was converted to pepsin (P) at pH 2.0, and the MW were determined as 32 kDa (for P-I), 31 kDa (for P-II) and 30 kDa (for P-III). The optimum pH and temperature of pepsins were 2.0-3.5, and 40-50 °C. All 3 pepsins were strongly inhibited by pepstatin A. Divalent cations slightly stimulated the pepsin activities, but ATP had no effect on the pepsins. Purified pepsins were effective in the hydrolysis of various proteins. Km and kcat of the three pepsins for hemoglobin hydrolysis were 107.64-276.61 µM and 18.30-32.68 s-1, respectively. The new pepsins have potential for use in protein food procession and modification.

Interactions of C. frondosa-derived inhibitory peptides against angiotensin I-converting enzyme (ACE), α-amylase and lipase.

The study illustrates the molecular mechanisms by which marine-derived peptides exhibited different structures and inhibition functions to concurrently inhibit multiple enzymes involved in chronic diseases. Peptides (2 mg/mL) exhibited inhibition against angiotensin-converting enzyme (ACE, inhibition of 52.2-78.8%), pancreatic α-amylase (16.3-27.2%) and lipase (5.3-17.0%). Further in silico analyses on physiochemistry, bioactivity, safety and interaction energy with target enzymes indicated that one peptide could inhibit multiple enzymes. Peptide FENLLEELK potent in inhibiting both ACE and α-amylase showed different mechanisms: it had ordered extended structure in ACE active pocket with conventional H-bond towards Arg522 which is the ligand for activator Cl-, while the peptide folded into compact "lariat" conformation within α-amylase active site and the K residue in peptide formed intensive H-bonds and electrostatic interactions with catalytic triad Asp197 - Asp300 - Glu233. Another peptide APFPLR showed different poses in inhibiting ACE, α-amylase and lipase, and it formed direct interactions to lipase catalytic residues Phe77 & His263.

Oxidative stress suppression in C. elegans by peptides from dogfish skin via regulation of transcription factors DAF-16 and HSF-1.

Functional peptides were obtained via enzymatic hydrolysis of smooth dogfish (Mustelus canis) skin. The enzyme-assisted process was optimized to achieve high yield of smooth dogfish skin peptides (SDSP). Fractions of SDSP (MW < 2 kDa, 2-5 kDa, 5-10 kDa and >10 kDa) showed in vitro antioxidant activities. The peptides <2 kDa (SDSP<2 kDa) significantly improved motility, reduced ROS and H2O2 levels of Caenorhabditis elegans, and increased its resistance to oxidative stress compared to the other peptide fractions. In vivo function of SDSP<2 kDa could be explained by their capacity to increase the expression of stress-response genes. The enhanced resistance to oxidative stress mediated by SDSP<2 kDa was dependent on DAF-16 and HSF-1. The amino acid residues and sequences of SDSP<2 kDa were characterized and revealed a higher content of hydrophobic versus polar amino acid contents. This study (especially the in vivo investigation) explored new potent antioxidant peptides derived from dogfish skin.

Effect of lactic fermentation on soy protein digestive pattern assessed by an in vitro dynamic gastrointestinal digestion model and the influence on human faecal microbiota.

BACKGROUND: The aim of this study was to investigate the effect of lactic fermentation on soy protein gastrointestinal digestive pattern and the influence of protein digesta on human faecal microbiota. Soymilk and soy yogurt were prepared in this study and a novel in vitro dynamic gastrointestinal model was employed to simulate gastric and duodenum digestions. Particle size, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), and peptide content were monitored at the end of duodenum tract. RESULTS: Ingestion of soy yogurt allowed a rapid drop in pH from 7.0 to 5.0 at simulated duodenal digestion (0-30 min), and resulted in a loss in soluble protein content compared to that of soymilk. The electrophoretic pattern between soymilk and soy yogurt exerted distinctive differences at early stages of duodenal digestion (0-60 min) and resulted in different peptide contents (180 min). Soy yogurt duodenal digesta collected at 180 min (D180), by co-fermentation with human intestinal flora distribution, allowed a higher population in Bifidobacterium spp., Lactobacillus/Enterococcus spp. and Streptococcus/Lactococcus spp., whereas soy yogurt D30 resulted in lower population in Clostridium and Escherichia coli compared to samples co-fermented with soymilk digesta. CONCLUSION: The results demonstrated lactic fermentation of soy protein modulated human intestinal microflora and might relate to the different protein digestive behaviours. © 2020 Society of Chemical Industry.

Preparation and physicochemical characterization of films prepared with salmon skin gelatin extracted by a trypsin-aided process.

The recovery of gelatins from Atlantic salmon (Salmo salar) skin for film formation and characterization was studied. Fish skins pre-treated with trypsin (250 U/g) produced the highest hydroxyproline content (7.41 ± 0.49 mg hydroxyproline/g treated skin) and yield (53.05 ± 4.38%) of gelatin, as compared to the use of saline solution. Pre-treatment with a lower concentration of trypsin (1 U/g) at a shorter pre-treatment time successfully reduced the degradation of gelatin with co-production of high molecular weight α-chains. Gelatin was further extracted by a trypsin-aided process for film formation and characterization. Films with increasing protein concentration (from 1 to 5%, w/v) exhibited higher thickness, tensile strength, and elongation at break (EAB), but a marked decrease in EAB for films with 6 and 7% (w/v). Films with 5% proteins showed higher thickness, lower tensile strength and higher EAB with increasing concentrations of glycerol (from 10 to 50% of proteins, w/w). All films exhibited high water uptake, decrease in light transmission and an increase in opacity as the protein and glycerol contents increased. Electrophoretic studies showed that the increase in the mechanical properties of the films was correlated with the increase in protein concentration, owing to the increased content of high molecular weight chain fractions. Furthermore, Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM) revealed the interaction between the proteins and glycerol for all films. This study demonstrated the viability of the trypsin supplementation process to obtain salmon skin gelatin for film formation.

Generation of antioxidative peptides from Atlantic sea cucumber using alcalase versus trypsin: In vitro activity, de novo sequencing, and in silico docking for in vivo function prediction.

A comprehensive evaluation was conducted to compare the generation of antioxidative peptides produced by alcalase versus trypsin from Atlantic sea cucumber. The in vitro antioxidative peptides were sequenced by de novo sequencing using LC-MS/MS. Key constituent antioxidative amino acids (KCAAA), i.e., Cys, His, Met, Trp and Tyr in the peptides and the molecular interactions between peptides and myeloperoxidase (MPO, a mediator and marker of in vivo oxidative stress), were analyzed by in silico methods. Alcalase-produced protein hydrolysates showed 5-35% higher in vitro antioxidant activity than the trypsin-produced ones. UPLC analysis revealed the total amino acid composition in peptide fractions <2 kDa. Alcalase produced 35.4% of peptides with both KCAAA and potential MPO inhibitory activity, compared with only 30.3% for trypsin. A representative peptide sequence TEFHLL generated by alcalase had intense molecular interactions with MPO active site, predicting a capacity to inhibit in vivo oxidative stress.

Optimal immobilization of trypsin from the spleen of albacore tuna (Thunnus alalunga) and its characterization.

Trypsin purified from the spleen of albacore tuna was immobilized onto Octyl Sepharose CL-4B, glutaraldehyde activated silica and 5'-4,4'-dimethyltryptamine-thymidine-succinyl controlled pore glass. Trypsin was highly and efficiently immobilized onto Octyl Sepharose CL-4B, with the highest activity (6.26 U/g support) and specific activity (1.45 U/mg bound protein). The optimum conditions for trypsin immobilization onto Octyl Sepharose CL-4B were 40 mg/mL trypsin solution, pH 7 at 4 °C for 6 h of incubation time. The optimal temperature and pH for the hydrolysis of N-α-benzoyl-DL-arginine-p-nitroanilide (DL-BAPNA) by the immobilized trypsin were 55 °C and 8.5, both of which were higher than that of the free form. In comparison with free enzyme, the immobilized trypsin exhibited greater resistances against thermal inactivation and organic solvents. The immobilized enzyme was less sensitive to inhibition by the soybean trypsin inhibitor compared with the free soluble form of the enzyme. According to the results, the immobilized trypsin and free enzyme retained 83% and 47% of their activity, respectively, when they were incubated with 1 µM of the soybean trypsin inhibitor. For the reusability study, the immobilized trypsin maintained 60% of its activity after 4 periods of activity, indicating that the immobilized trypsin had appropriate stability and could be reused.

Food-related transglutaminase obtained from fish/shellfish.

In the past two decades, there has been an upsurge of interest in transglutaminase (TGase) research due to their immense value in food applications to improve the quality as well as functional and nutritional attributes of food. Besides, TGases have been involved in new biomaterial development and shown to have potential in various biomedical applications. Presently, TGases from microbial sources (and some animal sources) are the main forms of the enzyme used in industrial applications. Nonetheless, there are disadvantages with the commercial TGases currently in use for food processing, such as the low activity, low yield, high cost, consumer aversions on their safety, and difficult in activity recovery, among others. Fish and shellfish TGases are promising alternatives for use in food industries due to their cheap sources, high yield and special characteristics such as cold activity and thermal-lability. This review presents at the onset, the catalytic mechanism of TGase based on updated research; compares the enzymatic properties of fish/shellfish TGases with their animal, plant and microbial counterparts; summarizes the unique properties of fish/shellfish TGases related with food usage; and discusses the current and potential applications of fish/shellfish TGases in foods.

Effects of different satiety levels on the fate of soymilk protein in gastrointestinal digestion and antigenicity assessed by an in vitro dynamic gastrointestinal model.

This study was designed to compare the effects of different satiety levels on the digestion and antigenicity of soymilk protein. The DIVRSD-II dynamic in vitro digestion model (DIVD) was employed to simulate different satiety degrees by changing the amount of food intake, namely full satiety (DIVD-FS), semi-satiety (DIVD-SS) and limited-satiety (DIVD-LS). A standardized static in vitro digestion method (SIVD) was used as a reference. Coupled with 60 min of gastric digestion and 120 min of intestinal digestion, the pH, particle size, soluble protein content, peptide content, gel electrophoresis, and antigenicity of soymilk digesta were monitored. The results showed that different satiety degrees altered soymilk protein digestive patterns in terms of gastric transit time, hydrolysis degree and antigenicity. The results of soluble protein content suggested that soymilk protein in DIVD-FS showed faster gastric transition than DIVD-SS and DIVD-LS. Gel electrophoresis and peptide content indicated that DIVD-FS digesta showed predominantly lower hydrolysis degrees in gastric and early intestinal stages. This further led to the persistence of major allergens at the early stage of intestinal digestion for DIVD-FS digesta. Soymilk protein digested using SIVD showed mostly intermediate values which indicated a good reference to the different satiety degrees of DIVD, although a descriptive result was obtained at early intestinal digestion. The results of this study showed for the first time the effect of different satiety degrees on hydrolysis kinetics, digestive degree and antigenicity of soy protein. The results suggested that the knowledge-base of soy protein digestibility depends on different satiety degrees and will be useful for guiding a healthy diet mode.

Preparation of Quercetin Loaded Microparticles and their Antitumor Activity against Human Lung Cancer Cells (A549) in vitro.

BACKGROUND: Novel quercetin-loaded microparticles (QM) were fabricated using coaxial electrospraying, characterized for surface morphology and release profile, and evaluated for antitumor activity in vitro. METHODS: QM exhibited an average diameter of 1.69 ±1.13 mm, which was an appropriate size suitable for respiratory delivery. X-ray diffraction patterns showed that the components in QM existed in an amorphous physical form, leading to favorable interactions between the drug (quercetin), the polymer matrix (polyvinylpyrrolidone, PVP) and other excipients (sodium dodecyl sulfate and sucralose). RESULTS: QM performed much faster release rate compared with free quercetin powder (Q) in vitro. Furthermore, QM also showed more potent inhibitory effects on A549 cell growth with reduced cell viability, decreased cell migration and induced more G0/G1 phase cell cycle arrest than Q. CONCLUSION: Thus, the quercetin loaded microparticles exhibited more potent inhibitory effects than free quercetin on A549 cell. The increased antitumor activity could be attributed to the enhanced accumulation of quercetin in the A549 cells with the QM. However, further studies are necessary to elucidate the exact mechanisms.

Anionic trypsin from the spleen of albacore tuna (Thunnus alalunga): Purification, biochemical properties and its application for proteolytic degradation of fish muscle.

Anionic trypsin from albacore tuna spleen was purified by chromatographic separations on Q-Sepharose, Superdex 75 and Arginine Sepharose 4B. The trypsin migrated as single bands in both SDS-PAGE and native-PAGE. The molecular weight of purified trypsin was estimated to be 30 kDa using SDS-PAGE. The enzyme exhibited maximal activity at pH 9.0 and 55 °C for hydrolysis of Boc-Val-Pro-Arg-MCA. pH and temperature stabilities of the trypsin were well maintained in the pH range of 6-11 and over a temperature range from 20 up to 50 °C. The enzyme was effectively inhibited by soybean trypsin inhibitor, N­tosyl­l­phenyl­alanine chloromethyl ketone (TLCK) and Pefabloc SC. The N-terminal amino acid sequence of 20 residues of the purified enzyme was IVGGYECQAHSQPHQVSLNA, which is highly homologous to other fish trypsins. The kcat/Km of the enzyme for Boc-Val-Pro-Arg-MCA was 2.60 ±â€¯0.07 s-1 mM-1. Purified trypsin also hydrolysed fish muscle proteins, suggesting its effectiveness in degradation of food proteins.

Health Insurance Portability and Accountability Act Noncompliance in Patient Photograph Management in Plastic Surgery.

BACKGROUND: Today, plastic surgeons have largely transitioned to digital photography. This shift has introduced new risks to daily workflows, notably data theft and Health Insurance Portability and Accountability Act (HIPAA) violations. METHODS: We performed a national survey of digital photograph management patterns among members of the American Society of Plastic Surgery and trainees in Accreditation Council for Graduate Medical Education-accredited plastic surgery programs. RESULTS: Our findings showed that attendings preferred the use of stand-alone digital cameras (91.4%), whereas trainees preferred the use of smartphones (96.1%) for capturing patient photographs. The rate of noncompliance was nearly identical; 82.8% of attendings were HIPAA noncompliant when using stand-alone digital cameras compared with 90.2% of trainees using smartphones. Both groups also breached HIPAA rules when using other photographic management modalities. CONCLUSIONS: This is the first study to quantify the prevalence of noncompliance with regard to an entire digital photograph management workflow. These findings were consistent with previous studies that reported that younger physicians tend to embrace newer technologies, whereas older attendings are more reluctant. The findings also suggest that HIPAA noncompliance in digital photograph security and management is a significant problem within the plastic surgery community.