Osteoinduction Using a Prefabricated Scaffold in the Gluteus Muscle of Wistar Rats: An Animal Study.

Introduction: Bone grafts are very significant for the reconstruction of the trauma and the deformations created thereafter. Various bone forming and promoting agents are widely studied. Hence, in this study, we aim to evaluate the osteoinduction effect of scaffold impregnated with Simvastatin in wistor rats in an observational study. Material and Methods: We implanted 18 Wistar rats to be equally divided as controls and test groups. Polycaprolactone coated with 20 mg concentration of Simavastatin under sterile condition was used as test and hydroxy appetite as control. Then, we observed the histopathological Heamatoxilin and Eosin and ABH, sections at 4, 12, and 26 weeks. We noted the calcium deposition and bone formation. Results: We observed a significant variation among groups at 4, 12, and 26 weeks in the calcium deposition. While for the bone formation no significant variation were noted among groups at 4 and 12 weeks, however, there was a significant difference noted at 26 weeks. At the end of 12 weeks, mild fibroblast proliferation was seen in the surrounding area. After 26 weeks, ABH sections showed focal area of fibroblast proliferation with mild mononuclear infiltration was noticed; but implant could not be located. Conclusion: We conclude that Simavastatin showed an anabolic effect on bone healing.

Preparation and optimization of chitosan nanoparticles from discarded squilla (Carinosquilla multicarinata) shells for the delivery of anti-inflammatory drug: Diclofenac.

Biological waste from marine sources is discarded into various water bodies which leads to dramatic increase in the water pollution near coastal areas. This animal waste consists of bioactive compounds such as fatty acids, amino acids, and chitin which can be used in agricultural and pharmaceutical sectors. The aim of the current study was to extract chitosan (CS) from the discarded shells of Carinosquilla multicarinata and prepare anti-inflammatory drug diclofenac potassium (DP) encapsulated chitosan nanoparticles (DP-CSNPs). The CS was extracted, purified and physicochemical and morphological properties were characterized such as viscosity (1.44cPs), molecular weight (~57 kDa), degree of deacetylation (83%). The DP-CSNPs were prepared by ionic gelation of extracted chitosan with tripolyphosphate (TPP) anions by varying chitosan, TPP, and drug concentrations. SEM imaging showed that DP-CSNPs were nano-sized (248 nm) along with small, spherical, and uniformity in shape. The endothermic peak appeared at 180°C while performing the thermal analysis of DP-CSNPs by differential scanning calorimetry (DSC). The Loading capacity (LC) and encapsulation efficiency (EE) were determined for all combinations while maximum EE (79.42%), LC (42.08%), and +0.00459 mV for Zeta potential were found for nanoparticles synthesized from CS with 2.5mg/mL concentration and 1mg/mL of TPP and drug concentrations. Moreover, in vitro drug release study was performed at simulated biological fluid (pH 7.4) and at 10th hr maximum (80%) of the drug was released from DP-CSNPs. Therefore, this waste source would be a better model system for the drug release. Implications: Dumping of marine waste into deep ocean has led to dramatic increase in water pollution leading to the endangerment of various oceanic animals. This discarded waste can be used sustainably for the isolation of various biopolymers into the ultimate use for human community. The work provides a detailed guide into the method of extraction of low molecular weight chitosan and preparation of nanoparticles for the delivery of anti-inflammatory drug diclofenac.

Radical scavenging and amino acid profiling of wedge clam, Donax cuneatus (Linnaeus) protein hydrolysates.

Body, foot and viscera of Donax cuneatus (Linnaeus) were hydrolyzed using commercial proteases (pepsin, trypsin and papain) and tested for their antioxidant activity by DPPH scavenging ability and reducing power assays. In comparison between all the hydrolysates, papain viscera (28.513 ± 0.165 & 0.186 ± 0.008) and foot (33.567 ± 0.132 & 0.166 ± 0.013) hydrolysates showed highest DPPH and reducing power ability respectively. The active hydrolysates were purified with DEAE- cellulose followed by Sephadex G-25 columns connected to FPLC. Further, the isolated active fractions were loaded onto HPLC for their amino acid profiling and found with the presence of potential amino acids viz., histidine, cysteine, alanine etc. These results suggest that the isolated antioxidant peptide from viscera and foot hydrolysate of D. cuneatus can be used in treating human diseases where free radicals and oxidative damage are involved.

Detection of collagen through FTIR and HPLC from the body and foot of Donax cuneatus Linnaeus, 1758.

To make more effective use of available marine resources, acid soluble collagen (ASC) was isolated from body and foot of wedge clam Donax cuneatus Linnaeus, 1758 with acetic acid and was characterized for their potential and commercial applications. The yield of ASC was 17% and 23% respectively. SDS PAGE, UV and FTIR spectroscopy showed that both were type I mainly with slight differences. HPLC was used for identifying the presence of different types of amino acids, where glycine was more or less 20% in both the samples and takes the lead amino acid position and presence of imino acids (11.8 and 12.6%) has been the characteristic feature of type I collagen.

Isolation of antioxidant peptides from clam, Meretrix casta (Chemnitz).

The antioxidant activity of marine clam, Meretrix casta (Chemnitz) protein hydrolysates prepared from different organs (body, foot and viscera), using the commercial enzymes (pepsin, trypsin and papain) were determined. The protein hydrolysate had a high antioxidant activity where, pepsin hydrolysate of viscera and trypsin hydrolysate of body and foot showed good activity. The viscera pepsin hydrolysate and foot trypsin hydrolysates were purified using FPLC on ion exchange and gel filtration chromatography procedure and activity was determined by DPPH radical scavenging and reducing ability assays. Further the amino acid content of the purified fractions was analyzed using HPLC. Active fractions contained good quantity of both essential and non-essential amino acids.

Antioxidant and functional properties of protein hydrolysates from pink perch (Nemipterus japonicus) muscle.

Functional properties and antioxidant activity of pink perch (Nemipterus japonicus) muscle hydrolysed by three different enzymes papain, pepsin and trypsin were studied. The protein hydrolysates produced by trypsin had an excellent solubility (98%) compared to pepsin (77%) and papain hydrolysate (74%). Conversely, the emulsifying activity index (ESI) and foaming abilities were affected by pH. DPPH radical scavenging ability, reducing power and metal chelating activity of protein hydrolysates increased with increase in concentration. Lipid peroxidation was strongly inhibited by 64% by protein hydrolysates produced by trypsin. The results revealed that the functional properties and antioxidant activities of pink perch were greatly affected by the enzymes used.

In vitro and in vivo studies on the antioxidant activity of fish peptide isolated from the croaker (Otolithes ruber) muscle protein hydrolysate.

Peptide from croaker (Otolithes ruber) muscle protein hydrolysate was purified, characterized and evaluated for its in vitro and in vivo antioxidant activity. Results showed that purified peptide contained the amino acid sequence as Lys-Thr-Phe-Cys-Gly-Arg-His (861.6Da), which were expected to contribute to its antioxidant activities. This peptide efficiently quenched 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radicals (84.5±1.2 and 62.4±2.9%), and successfully inhibits the lipid peroxidation and DNA damage and proven to be a potent antioxidant at different in vitro systems. It also improved the endogenous cellular antioxidant enzymes in Wistar rat by increasing the activities of catalase (CAT), glutathione-S-transferase (GST) and superoxide dismutase (SOD) after supplementation of the peptide (283.6±7.25, 4.3±0.78 and 28.42±1.97) compared to the negative control (196.4±5.65, 1.3±0.45 and 15.1±0.35). Therefore, croaker muscle peptide can increase an endurance capacity and facilitate recovery from oxidative stress.

Purification and identification of antioxidant peptides from the skin protein hydrolysate of two marine fishes, horse mackerel (Magalaspis cordyla) and croaker (Otolithes ruber).

In the current study, two peptides with antioxidant properties were purified from skin protein hydrolysates of horse mackerel (Magalaspis cordyla) and croaker (Otolithes ruber) by consecutive chromatographic fractionations including ion exchange chromatography and gel filtration chromatography. By electron spray ionization double mass spectrometry (ESI-MS/MS), the sequence of the peptide from the skin protein hydrolysate of horse mackerel was identified to be Asn-His-Arg-Tyr-Asp-Arg (856 Da) and that of croaker to be Gly-Asn-Arg-Gly-Phe-Ala-Cys-Arg-His-Ala (1101.5 Da). The antioxidant activity of these peptides was tested by electron spin resonance (ESR) spectrometry using 1-diphenyl-2-picryl hydrazyl (DPPH·) and hydroxyl (OH·) radical scavenging assays. Both peptides exhibited higher activity against polyunsaturated fatty acid (PUFA) peroxidation than the natural antioxidant α-tocopherol. These results suggest that the two peptides isolated from the skin protein hydrolysates of horse mackerel and croaker are potent antioxidants and may be effectively used as food additives and as pharmaceutical agents.

Optimization of enzymatic hydrolysis conditions for the production of antioxidant peptides from muscles of Nemipterus japonicus and Exocoetus volitans using response surface methodology.

In the present study, protein of muscles of commercially important marine fishes Nemipterus japonicus and Exocoetus volitans were extracted by trypsin and their hydrolysis conditions viz., temperature, time, and enzyme to substrate concentration on degree of hydrolysis were studied by response surface methodology. The optimum values for N. japonicus was found as temperature, 30°C, hydrolysis time of 100 min an enzyme/substrate concentration of 1.59% whereas, for E. volitans muscle protein, optimum hydrolysis conditions were temperature, 30°C, hydrolysis time of 115 min and enzyme/substrate concentration of 1.67%. Furthermore, amino acid sequence of antioxidant peptides derived after chromatographic purification was identified by ESI-MS/MS. The analysis of peptides showed sequences as Glu-Ser-Asp-Arg-Pro (620.3 Da) and Gly-Trp-Met-Gly-Cys-Trp (747.3) for N. japonicus and E. volitans muscle, respectively. The peptides contained important antioxidant amino acids and acted as good antioxidant peptides to scavenge free radicals.

Anticoagulant, antiherpetic and antibacterial activities of sulphated polysaccharide from Indian medicinal plant Tridax procumbens L. (Asteraceae).

The sulphated polysaccharide from the widespread Tridax procumbens plant was studied for the anticoagulant, antiherpetic and antibacterial activity. The anticoagulant activity was determined by the activated partial thromboplastin time assay. The sulphated polysaccharide from T. procumbens represented potent anticoagulant reaching the efficacy to heparin and chondroitin sulphate. Moreover, the sulphated polysaccharide extracted from T. procumbens was found non-toxic on Vero cell lines up to the concentration of 200 µg/ml. Sulphated polysaccharide exhibited detectable antiviral effect towards HSV-1 with IC(50) value 100-150 µg/ml. Furthermore, sulphated polysaccharide from T. procumbens was highly inhibitory against the bacterial strains Vibrio alginolyticus and Vibrio harveyi isolated from oil sardine.

Purification and biochemical characterization of antioxidant peptide from horse mackerel (Magalaspis cordyla) viscera protein.

In the present study, a peptide having high antioxidant properties was isolated from horse mackerel viscera protein, Magalaspis cordyla. In vitro gastrointestinal digestion was employed to obtain potential protein hydrolysate and was subjected to consecutive chromatographic methods using fast protein liquid chromatography (FPLC) connected to diethyl amino ethyl (DEAE) anion exchange column and Sephadex G-25 gel filtration column. The activity of the fractions was tested against DPPH and hydroxyl radicals and the isolated peptide showed 89.2 and 59.1 percentage of scavenging. The amino acid sequence of purified peptide was determined using ESI-MS/MS as Ala-Cys-Phe-Leu (518.5 Da), it exhibited high activity against polyunsaturated fatty acid (PUFA) peroxidation than that of natural antioxidant, α-tocopherol.

American cutaneous leishmaniasis in Guyana, South America.

The 185 patients who presented at the dermatology clinic of Georgetown Public Hospital, Guyana, between 1992 and 1998, with skin ulcers indicative of American cutaneous leishmaniasis (ACL) were retrospectively reviewed. The laboratory-confirmed cases of ACL were identified and the corresponding data were analysed for risk factors such as age, gender, areas of residence and of possible exposure to the causative agent (Leishmania braziliensis guyanensis), ethnic origin, longevity of the ulcers, and treatment regimes prior to the definitive diagnosis. Eighty-one (43%) of the 185 subjects were confirmed to be infected with Le. b. guyanensis. Although 53 (66%) of the cases lived in or close to the capital city, Georgetown, most of the cases had travelled to (and probably been infected in) region X in the interior of Guyana (32%) or regions VII (23%), VIII (23%), IX (11%), VI (5%), I (3%) or III (3%), usually because they were involved in the mining (41%) or lumber (21%) industries, the army or hunting. Almost all (95%) of the cases were male and most (58%) were aged 20-39 years. In general, the cases had had their skin lesions for many days before presenting for treatment: 46% for 1-5 weeks and 3% for > 6 months. Prior to presentation at the clinic, many of the cases had attempted to cure themselves, using local herbal remedies (37%), antibiotics and antifungal remedies (39%), other creams (5%), household chemicals (9%) or miscellaneous remedies such as lead salts (especially lead sulphate) and battery acid, all without success. Recommendations are made for an epidemiological study of active ACL among forest workers, eco-tourists and residents of high-risk areas. Diagnostic centres need to be sited in the regions most at-risk, particularly in or near environments in which the main vectors - sandflies such as Lutzomyia umbratilis, Lu. anduzei and Lu. whitmani - are known to be prevalent.