Characterizations and fluoride adsorption performance of wattle humus biosorbent.

Considering the serious health effects of fluoride contamination, an environment friendly bioadsorbent was derived from wattle humus for fluoride removal by conventional thermal activation process. Analytical characterizations revealed that heterogeneous morphological textured wattle humus enabled remarkable adsorption capacity. XPS analysis substantiated that fluoride had been successfully adsorbed on to the carbonized wattle humus surface through chemisorption. Fluoride adsorption efficiency was systematically rationalized via batch adsorption studies. Experiments were performed at different initial fluoride concentration and scrutinized the impact of contact time (10-120 min), adsorbent dosage (0.5-2.5 g), pH (2.0-9.0), and interfering co-existing ions (SO42-, NO3-, Cl-, and HCO3-) on fluoride removal. Even at different adsorbate dosage (2-10 mg/L), 98% fluoride removal efficiency was achieved under pH > 6. The competitive anions do not interfere the wattle humus fluoride adsorption capacity. Moreover, the adsorption isotherms and kinetics studies inferred that monolayer and multilayer adsorption behavior by wattle humus leads to noticeable fluoride adsorption. Adsorbent regeneration test affirms that regenerated adsorbent found higher (>95%) fluoride removal efficiency even at five recycle runs.

Wattle fencing improved soil aggregate stability, organic carbon stocks and biochemical quality by restoring highly eroded mountain region soil.

Eroded bare land stabilization is important to reduce soil erosion and stimulate soil carbon (C) sequestration for improved soil biogeochemical quality in hillslope soils. This study investigated the effectiveness of wattle fencing as a bioengineering tool to improve soil stabilization, soil physico-chemical properties and soil organic C dynamics and reduce soil erodibility in the Boyabat mountain regions of Turkey with rough and over-steepened slope (50-70%). Wattle fence treatments were developed in the area of 50 ha in the spring season of 2010 and surface (0-20 cm) and subsurface soil (20-40 cm) samples were taken in Spring, 2015. Results revealed that, compared to control with bare slope, wattle fencing significantly improved some soil physico-chemical, and microbial properties and erodibility indices by increasing clay ratio, dispersion ratio and aggregate stability index in surface and subsoils. Wattle fencing enhanced plant available water contents more in surface than in subsoils. Wattle fencing also increased microbial biomass C contents by 55% and 43% in surface and subsurface soils, respectively. Soil organic C followed similar trends; however, they were indifferent between sampling depths for the control soils. Soil organic C stocks and aggregate stability index were significantly positively correlated and seemed to be better predictor of positive effects of wattle fencing on soil structural stability, erodibility and associated properties. We found positive effects of soil organic C contents on microbial biomass C and soil-water relations suggesting restoration of soil biological functions and favorable influence on soil water retention following wattle fencing. Although sparse vegetation was observed in the research area, our study emphasizes performing further research to understand the effects of wattle fencing along with afforestation with native vegetation on soil erosion rates on a long-term basis by considering the variability in edaphic and environmental factors.

Identification of Angiotensin I-Converting Enzyme-Inhibitory and Anticoagulant Peptides from Enzymatic Hydrolysates of Chicken Combs and Wattles.

Peptides from protein hydrolysate of a mixture of chicken combs and wattles (CCWs) were obtained through enzymatic hydrolysis, and their anticoagulant and inhibitory effects on angiotensin I-converting enzyme (ACE) were investigated. The protein hydrolysate exhibited anticoagulant capacity by the intrinsic pathway (activated partial thromboplastin time) and potent ACE-inhibitory activity. The peptides were sequenced by LC-MS to identify those with higher inhibitory potential. From the pool of sequenced peptides, the following three peptides were selected and synthesized based on their low molecular weight and the presence of amino acids with ACE-inhibitory potential at the C-terminus: peptide I (APGLPGPR), peptide II (Piro-GPPGPT), and peptide III (FPGPPGP). Peptide III (FPGPPGP) showed the highest ACE-inhibitory capacity among the peptides selected. In conclusion, a peptide (FPGPPGP) of unknown sequence was identified as having potent ACE-inhibitory capacity. This peptide originated from unconventional hydrolysates from poultry slaughter waste, including combs and wattles.

Effectiveness of a low-fat yoghurt supplemented with rooster comb extract on muscle strength in adults with mild knee pain and mechanisms of action on muscle regeneration.

PURPOSE: To determine the effects of the intake of low-fat yoghurt supplemented with rooster comb extract (RCE) on muscle strength. METHODS AND RESULTS: 148 subjects, with mild knee pain, participated in a randomized, placebo-controlled, double-blind, and parallel study. Muscle strength, knee effusion, and pain perception were measured. C2C12 myoblasts were used to elucidate the mechanisms of action involved. RCE improved total work and mean power in men, and also peak torque in extension by 10%. RCE reduced synovial effusion by 11.8% and pain perception by 24.6%. Both RCE and HA increased myoblast proliferation by 29%, while RCE reduced myoblast differentiation by 36.2%, suggesting a beneficial role of RCE in muscle regeneration. CONCLUSIONS: Low-fat yoghurt supplemented with RCE improved muscle strength. This effect is partially explained by muscle regeneration enhancement, reduced synovial effusion, and reduced pain perception, which could exert a beneficial clinical impact on men affected by mild knee pain.

A low-fat yoghurt supplemented with a rooster comb extract on muscle joint function in adults with mild knee pain: a randomized, double blind, parallel, placebo-controlled, clinical trial of efficacy.

Preliminary results suggested that oral-administration of rooster comb extract (RCE) rich in hyaluronic acid (HA) was associated with improved muscle strength. Following these promising results, the objective of the present study was to evaluate the effect of low-fat yoghurt supplemented with RCE rich in HA on muscle function in adults with mild knee pain; a symptom of early osteoarthritis. Participants (n = 40) received low-fat yoghurt (125 mL d(-1)) supplemented with 80 mg d(-1) of RCE and the placebo group (n = 40) consumed the same yoghurt without the RCE, in a randomized, controlled, double-blind, parallel trial over 12 weeks. Using an isokinetic dynamometer (Biodex System 4), RCE consumption, compared to control, increased the affected knee peak torque, total work and mean power at 180° s(-1), at least 11% in men (p < 0.05) with no differences in women. No dietary differences were noted. These results suggest that long-term consumption of low-fat yoghurt supplemented with RCE could be a dietary tool to improve muscle strength in men, associated with possible clinical significance. However, further studies are needed to elucidate reasons for these sex difference responses observed, and may provide further insight into muscle function.

Proteolytic activity from chicken intestine and pancreas: extraction, partial characterization and application for hyaluronic acid separation from chicken comb.

BACKGROUND: The potential utilization of viscera and combs, by-products from the chicken-processing industry, was investigated. Chicken combs have been reported to consist of hyaluronic acid (HA) bound to protein. Thus proteases extracted from broiler viscera were applied to digest combs to separate HA, a high-value ingredient widely used in pharmaceutical and medical products. RESULTS: The highest activity of crude protease extracted from chicken intestine and pancreas was 0.35 U mg(-1) enzyme at 60 °C and pH 7.5. pH stability of the enzyme was attained from pH 6 to 8, while its thermal stability declined from 30 to 50 °C, with complete activity loss occurring after 30 min at temperatures above 60 °C. Therefore the optimal conditions for broiler comb digestion by crude protease in this study were chosen as pH 7.5 and 50 °C. The results showed that the yield of crude enzyme-extracted HA was lower (P < 0.05) than that obtained by commercial papain digestion. Similar identity of extracted HA and HA standard was verified by cellulose acetate electrophoresis and Fourier transform infrared spectroscopy. CONCLUSION: The crude enzyme extract from chicken intestine and pancreas had high proteolytic activity and could be used successfully to separate HA from broiler combs. The method provides an appropriate means to add value to poultry-processing waste.

Genotoxicity, acute and subchronic toxicity studies in rats of a rooster comb extract rich in sodium hyaluronate.

The toxicity of a rooster comb extract (IB0004) that contains mainly sodium hyaluronate was assessed in acute and subchronic studies and in a bacterial reverse mutation assay. In a single dose acute study, male and female rats were administered 2000 mg/kg body weight (bw) of the product and observed for 14 days. No mortality was recorded, thus it was considered that the minimum lethal dose for rats by oral route was greater than 2000 mg/kg bw. A 90-day subchronic study (5, 55 and 600 mg/kg bw/day, oral gavage) with 50 male and 50 female Wistar-Hannover rats produced no significant adverse effects on food consumption, body weight, mortality, clinical biochemistry, hematology, gross pathology, and histopathology. Although some differences were observed between the treated and control animals in body weight gain (%) and some hematological parameters, these changes were generally minor in nature and, are considered to be of no toxicological significance. The no-observable-adverse-effects level was established at 600 mg/kg bw/day. There was no evidence of mutagenic activity in Salmonella typhimurium TA98, TA100, TA1535 and TA1537 or in Escherichia coli WP2 uvra pkM101. In conclusion, the results from these safety studies support the safety of rooster comb extract IB0004 in food.

Extraction of hyaluronic acid (HA) from rooster comb and characterization using flow field-flow fractionation (FlFFF) coupled with multiangle light scattering (MALS).

Hyaluronic acid (HA) was extracted in a relatively large scale from rooster comb using a method similar to that reported previously. The extraction method was modified to simplify and to reduce time and cost in order to accommodate a large-scale extraction. Five hundred grams of frozen rooster combs yielded about 500 mg of dried HA. Extracted HA was characterized using asymmetrical flow field-flow fractionation (AsFlFFF) coupled online to a multiangle light scattering detector and a refractive index detector to determine the molecular size, molecular weight (MW) distribution, and molecular conformation of HA. For characterization of HA, AsFlFFF was operated by a simplified two-step procedure, instead of the conventional three-step procedure, where the first two steps (sample loading and focusing) were combined into one to avoid the adsorption of viscous HA onto the channel membrane. The simplified two-step AsFlFFF yielded reasonably good separations of HA molecules based on their MWs. The weight average MW (M(w) ) and the average root-mean-square (RMS) radius of HA extracted from rooster comb were 1.20×10(6) and 94.7 nm, respectively. When the sample solution was filtered through a 0.45 µm disposable syringe filter, they were reduced down to 3.8×10(5) and 50.1 nm, respectively.

Effect of a natural extract of chicken combs with a high content of hyaluronic acid (Hyal-Joint) on pain relief and quality of life in subjects with knee osteoarthritis: a pilot randomized double-blind placebo-controlled trial.

BACKGROUND: Intra-articular hyaluronic acid represents a substantive addition to the therapeutic armamentarium in knee osteoarthritis. We examined the effect of dietary supplementation with a natural extract of chicken combs with a high content of hyaluronic acid (60%) (Hyal-Joint) (active test product, AP) on pain and quality of life in subjects with osteoarthritis of the knee. METHODS: Twenty subjects aged > or =40 years with knee osteoarthritis (pain for at least 15 days in the previous month, symptoms present for > or =6 months, Kellgren/Lawrence score > or =2) participated in a randomized double-blind controlled trial. Ten subjects received AP (80 mg/day) and 10 placebo for 8 weeks. The Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) and quality of life by the Short Form-36 (SF-36v2) were administered at baseline and after 4 and 8 weeks of treatment. RESULTS: WOMAC pain (primary efficacy variable) was similar in both study groups (mean [SD]) with 6.6 (4.0) points in the AP group and 6.4 (2.7) in the placebo group (P = 0.943). As compared with baseline, subjects in both groups showed statistically significant improvements in WOMAC pain, stiffness, physical function subscales, and in the aggregate score, but the magnitude of changes was higher in the AP group for WOMAC physical function (-13.1 [12.0] vs. -10.1 [8.6], P = 0.575) and total symptoms (-18.6 [16.8] vs. -15.8 [11.4], P = 0.694). At 4 weeks, statistically significant mean changes compared with baseline were observed in the SF-36v2 scales of role-physical, bodily pain, social functioning and role-emotional among subjects in the AP group, and in physical functioning, bodily pain, and social functioning in the placebo group. At 8 weeks, changes were significant for role-physical, bodily pain, and physical component summary in the AP group, and for physical functioning and role-emotional in the placebo arm. Changes in bodily pain and social functioning were of greater magnitude in subjects given AP. CONCLUSION: This pilot clinical trial showed that daily supplementation with oral hyaluronic acid from a natural extract of chicken combs (Hyal-Joint) was useful to enhance several markers of quality of life in adults with osteoarthritis of the knee. The results warrant further study in larger sample sizes.

Intra-articular injection of hyaluronate (SI-6601D) improves joint pain and synovial fluid prostaglandin E2 levels in rheumatoid arthritis: a multicenter clinical trial.

OBJECTIVE: The relationship between clinicalfeatures and biochemical parameters of synovialfluid after serial intra-articular injections of sodium hyaluronate (SI-6601D) was investigated. METHODS: SI-6601D (sodium hyaluronate with an average molecular weight of 8.4 x 10(5); 25mg/2.5ml/syringe) was injected intra-articularly into the knees of 25 patients with rheumatoid arthritis (RA) every week for 5 consecutive weeks. Clinical and biochemical parameters were monitored before and after injection. Clinicalfindings included pain, as a summation of 3 categories (pain at rest, pain in motion and pain in passive motion, each assessed on a 4-step rating scale), and inflammation, also as a summation of 3 categories (swelling, patellar ballotement and local warmth, each assessed on a 4-step rating scale). Pain on walking of patient was qualitatively assessed by visual analogue scale (VAS). The aspirated volume of synovialfluid (SFV) was recorded and levels of prostaglandin (PG) E2, transforming growth factor beta-1, tumor necrosis factor alpha, interleukin I receptor antagonist, chondroitin 4-sulfate (C4S) and chondroitin 6-sulfate were measured. RESULTS: Significant improvement in pain symptoms (p < 0.0001), inflammation (p < 0.0001), VAS pain (p < 0.001) and SFV (p < 0.05) were observed after the 5 injections. Levels of PGE2 (p < 0.05) and C4S (p < 0.05) in the synovialfluid were significantly decreased. DISCUSSION: SI-6601D improved local clinical symptoms in RA patients by suppressing PGE2 and, therefore, may be a useful treatment for local inflammation in RA.

Single-step divergent selection for male comb shape in two White Leghorn lines.

1. Divergent selection for comb shape (SH, the way that the cockerel bears the comb) was performed in 2 White Leghorn lines in a study aimed at assessing possibilities for improving SH. 2. Line A, selected for large comb size (CS) at 29 weeks of age, had great SH problems whereas Line H, selected for high hyaluronic acid concentration in the comb (HA), had minor SH problems. Multivariate analyses were used to estimate genetic parameters for SH, CS and HA in lines A and H and in a control line (C). 3. Significant direct selection responses for SH were achieved in both lines. There were significant unfavourable correlated responses for CS in both lines. 4. The correlated responses for HA were not significant and were unfavourable in Line A and favourable in Line H. 5. Heritabilities for SH and CS were high in both lines and relatively low for HA. Most of the genetic correlations were in agreement with the correlated responses obtained.

Interaction of hyaluronic acid with mucin, evaluated by gel permeation chromatography.

Hyaluronic acid (HA) is known to increase the ocular bioavailability of ophthalmic drugs not only for its viscous properties but also for its specific affinity for ocular mucins. This phenomenon, called bio- or mucoadhesion, can be evaluated in vitro by mechanical tests which, however, require considerable amounts of mucin (M) that are difficult to obtain from ocular surfaces. Thus, we developed an alternative method, based on gel permeation liquid chromatography, to examine the interaction of HA with microgram quantities of mucin. HA (from human umbilical cord or rooster comb) were fractionated using a Sepharose CL-4B column, before and after incubation with porcine gastric mucin (PGM), and the fractions were analyzed by a specific assay based on the histological dye Stains-all. PGM interacted with high molecular weight (M.W). HA, causing the displacement of low M.W., non-covalently bound, HA fragments, which were eluted under a distinct chromatographic peak. By quantitating the relative area of this peak, an evaluation of the mucoadhesion of HA could be obtained. This method could be useful to study the interaction between HA and microgram quantities of ocular M (mucin), obtained from individual patients or normal subjects.

Light microscopic histochemical and immunohistochemical localisation of sulphated glycosaminoglycans in the rooster comb and wattle tissues.

Comb and wattle tissues, which consist of layers of epidermis, dermis and central connective tissue, are known to contain sulphated glycosaminoglycans (GAGs) including dermatan sulphate and chondroitin sulphate-dermatan sulphate copolymers. Little is known about distribution of these CAGs in each tissue. The objective of this study was to localise sulphated GAGS in the comb and wattle tissues from mature roosters. Monoclonal antibodies 6D6, CS-56 and AH12 specific to dermatan sulphate proteoglycan (decorin), chondroitin sulphate and keratan sulphate, respectively, were used. In both tissues, 6D6 epitope was found to be more concentrated in the superficial layer of dermis and the central connective tissue than in the intermediate layer of dermis containing fibromucoid tissue. The staining pattern for 6D6 epitope was similar to that for collagen fibres. In contrast, CS-56 epitope was uniformly distributed in most parts of the dermis and the central connective tissue. The stratum germinativum in the epidermis was the major tissue showing positive staining with AH12, haematoxylin and safranin-O.

Rooster comb and wattle tissues contain an anti-keratan sulfate monoclonal antibody epitope.

Glycosaminoglycan fractions isolated from either comb or wattle tissue were examined using ELISA for their antigenicities to AH12, a monoclonal antibody recognizing keratan sulfate. The results showed a positive antibody binding to a sulfated glycosaminoglycan fraction from either tissue. The treatment of the same fraction with keratan sulfate degrading enzyme, keratanase, or endo-beta-galactosidase resulted in a decrease in its antigenicity. These findings indicated the presence of AH12 epitope (keratan sulfate disaccharide) in both tissues.

Research note: the reactivity of anti-chondroitin sulfate antibody to comb and wattle galactosaminoglycans.

The current study was undertaken to examine the reactivity of anti-chondroitin sulfate monoclonal antibody, CS-56, to the comb and wattle galactosaminoglycans containing varying proportions of L-iduronic acid. Galactosaminoglycans were isolated from mature rooster comb and wattle tissues and were separated into four fractions (I, II, III, and IV), which were precipitable with 18, 25, 40, and 50% ethanol, respectively. Each fraction was examined using an ELISA technique. The results demonstrated presence of CS-56 epitope in galactosaminoglycans from both tissues. Most (> 99%) of the epitope were found in Fractions III and IV, which were galactosaminoglycans rich in D-glucuronic acid. Fractions I and II, which were dermatan sulfates rich in L-iduronic acid, showed little or very weak antigenicities.

A quantitative chemical study of the comb and wattle galactosaminoglycans from single comb White Leghorn roosters.

Galactosaminoglycans from mature rooster comb and wattle tissues were separated into five fractions by ethanol precipitation. An average of 90% total uronic acid was recovered in Fractions I to III. Fractions I and II were dermatan sulfate with relatively high proportions of L-iduronic acid (61 to 80%), but this uronic acid was a minor component (30%) in Fraction III, in which D-glucuronic acid was the major uronic acid. Digestion with testicular hyaluronidase suggested that most if not all of the galactosaminoglycans in Fractions I to III were copolymers containing both L-iduronic acid and D-glucuronic acid. Fractions IV and V contained much lower proportions of L-iduronic acid and showed broader electrophoresis bands than did Fraction III.

Hyaluronic acid--syneretic glycosaminoglycan.

Hyaluronic acid from different sources, umbilical cord, vitreous, rooster comb and streptococcus, all exhibit a unique hydration behavior. Each hyaluronic acid and each different salt form has a maximum non-freezable water content at a set concentration beyond which the bound (non-freezable) water decreases. This type of behavior indicates a syneretic process simply due to concentration alone.

Chemical composition of glycosaminoglycan fractions from the comb and wattle of single comb white Leghorn roosters.

Glycosaminoglycan (GAG) fractions were isolated from papain digests of comb and wattle tissues of 52-wk-old Single Comb White Leghorn roosters and the chemical composition of each fraction was examined. Total GAG concentrations were greater in the comb than in the wattle tissue with the ratio of hyaluronic acid to sulfated GAG being similar between the two tissues. L-iduronosyl-N-acetylgalactosamine-4-sulfate was the predominant disaccharide unit in the sulfated GAG fraction.