Bile acid structure-activity relationship: evaluation of bile acid lipophilicity using 1-octanol/water partition coefficient and reverse phase HPLC.

Two independent methods have been developed and compared to determine the lipophilicity of a representative series of naturally occurring bile acids (BA) in relation to their struc- ture. The BA included cholic acid (CA), chenodeoxycholic acid (CDCA), ursodeoxycholic acid (UDCA), deoxycholic acid (DCA), hyodeoxycholic acid (HDCA), ursocholic acid (UCA), hyocholic acid (HCA), as well as their glycine and taurine ami- dates. Lipophilicity was determined using a 1-octanol/water shake-flask procedure and the experiments were performed at different pH and ionic strengths and at initial BA concentrations below their critical micellar concentrations (CMC) and the water solubility of the protonated form. The experimental data show that both the protonated (HA) and ionized (A-) forms of BA can distribute in 1-octanol, and consequently a partition co- efficient for HA (logP' HA) and for A- (logP' A-) must be defined. An equation to predict a weighted apparent distribution coefficient (D) value as a function of pH and pKa has been de- veloped and fits well with the experimental data. Differences be- tween logP for protonated and ionized species for unconjugated BA were in the order of 1 log unit, which increased to 2 for glycine-amidated BA. The partition coefficient of the A- form in- creased with Na+ concentration and total ionic strength, suggest- ing an ion-pair mechanism for its partition into 1-octanol. Lipophilicity was also assessed using reverse phase chromatogra- phy (C-18-HPLC), and a capacity factor (K') for ionized species was determined. Despite a broad correlation with the logP data, some BA behaved differently. The logP values showed that the order of lipophilicity was DCA >CDCA >UDCA > HDCA > HCA>CA >UCA for both the protonated and ionized uncon- jugated and glycine-amidated BA, while the K' data showed an inversion for some BA, i.e., DCA>CDCA >CA> HCA> UDCA > HDCA >UCA. The logP data fitted well with other in- direct measurements of BA monomeric lipophilicity such as al- bumin binding or accessible total hydrophobic surface area data calculated by energy minimization and molecular computer graphics. Differences between unconjugated and amidated BA are consistent with the presence of an amide bond and a lower pKa when pH dependence was studied. Capacity factors, on the other hand, were related to properties of BA micelles such as cholesterol-solubilizing capacity and membrane disruption, reflecting the BA detergency. The extrapolation of these data to biological phenomena must carefully consider the experimental conditions in which the interaction occurs, Le., total BA concen- tration, ionic strength, Na+ concentration, and pH, which in turn determine the BA species existing in solution that coul

Improvement of resveratrol permeation through sublingual mucosa: Chemical permeation enhancers versus spray drying technique to obtain fast-disintegrating sublingual mini-tablets

Resveratrol (RSV) is a natural polyphenol with several interesting broad-spectrum pharmacological properties. However, it is characterized by poor oral bioavailability, extensive first-pass effect metabolism and low stability. Indeed, RSV could benefit from the advantage of the sublingual route of administration. In this view, RSV attitudes to crossing the porcine sublingual mucosa were evaluated and promoted both by six different chemical permeation enhancers (CPEs) as well as by preparing four innovative fast-disintegrating sublingual mini-tablets by spray drying followed by direct compression. Since RSV by itself exhibits a low permeation aptitude, this could be significantly enhanced by the use of CPEs as well as by embedding RSV in a spray-dried powder to be compressed in order to prepare fast-disintegrating mini-tablets. The most promising observed CPEs (menthol, lysine and urea) were then inserted into the most promising spray-dried excipients’ compositions (RSV-B and RSV-C), thus preparing CPE-loaded mini-tablets. However, this procedure leads to unsatisfactory results which preclude the possibility of merging the two proposed approaches. Finally, the best spray-dried composition (RSV-B) was further evaluated by SEM, FTIR, XRD and disintegration as well as dissolution behavior to prove its effectiveness as a sublingual fast-disintegrating formulation

Influence of rainfall on E. coli concentrations in clams: results of collaboration between competent health authority and producers’ association in the Province of Fermo (Italy)

The Area Vasta di Fermo (the competent health authority of the Marche Region), in collaboration with the local Producers’ Association conducted a series of studies on the presence of E. coli as an indicator of faecal contamination in the environment, in shellfish and in fishery waters, and on the factors that can affect their presence. These studies, carried out from 2008 to 2011, included an assessment of the currents along the coast, of the precipitations, and data from the monitoring of E. coli on shellfish harvested in the collection areas were examined. The results showed that in most cases, small concentrations of microorganisms in shellfish corresponded to little or no precipitations, while an increase in the levels of E. coli was preceded by more or less abundant rainfalls. The conclusions suggest that it is advisable to carry out a more detailed risk analysis which should take into account the above-mentioned factors. Furthermore, monitoring alone based on the determination of indicator organisms, especially when carried out as a single analysis or with a small number of E. coli determinations, does not provide a satisfactory indication of safety. The regional surveillance plans should be applied timely and rigorously, together with on-site investigations aimed at identifying changes that can affect the presence of E. coli in shellfish. Food business operators themselves could implement good manufacturing practices to verify whether the microbiological parameters are within the prescribed limits after rainfalls, especially if heavy

Lipid nanocarriers-loaded nanocomposite as a suitable platform to release antibacterial and antioxidant agents for immediate dental implant placement restorative treatment

Immediate implant placement is a single-stage restorative approach for missing teeth widely used to overcome the ridge remodeling process occurring after dental extractions. The success of this procedure relies on opportune osseointegration in the surrounding tissues. To support this process, a multifunctional nanocomposite, to be applied in the fresh post-extraction socket, was here designed, prepared, and characterized. This formulation consists of quercetin (QRC)-loaded nanostructured lipid carriers (NLCs) entrapped in a chitosan-based solid matrix containing ciprofloxacin (CPX). QRC-NLCs were prepared by homogenization followed by high-frequency sonication, and thereafter this dispersion was trapped in a chitosan-based CPX-loaded gel, obtaining the nanocomposite powder (BioQ-CPX) by lyophilization. BioQ-CPX displayed desirable properties such as high porosity (94.1 ± 0.5%), drug amounts (2.1% QRC and 3.5% CPX). and low swelling index (100%). Moreover, the mechanism of drug release from BioQ-CPX and their ability to be accumulated in the target tissue were in vitro and ex vivo elucidated, also by applying mathematical models. When trapped into the nanocomposite, QRC stressed under UV light exposure (50 W) was shown to maintain its antioxidant power, and CPX and QRC under natural light were stable over nine months. Finally, both the measured antioxidant power and the antimicrobial and antibiofilm properties on Staphylococcus aureus demonstrated that BioQ-CPX could be a promising platform to support the single-stage dental restorative treatment

PRESENCE OF TRICHINELLA BRITOVI IN WILD BOAR IN THE MARCHE REGION REGULARLY SLAUGHTERED

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Novel splice-affecting variants in CYP27A1 gene in two Chilean patients with Cerebrotendinous Xanthomatosis

Cerebrotendinous Xanthomatosis (CTX), a rare lipid storage disorder, is caused by recessive loss-of-function mutations of the 27-sterol hydroxylase (CYP27A1), producing an alteration of the synthesis of bile acids, with an accumulation of cholestanol. Clinical characteristics include juvenile cataracts, diarrhea, tendon xanthomas, cognitive impairment and other neurological manifestations. Early diagnosis is critical, because treatment with chenodeoxycholic acid may prevent neurological damage. We studied the CYP27A1 gene in two Chilean CTX patients by sequencing its nine exons, exon-intron boundaries, and cDNA from peripheral blood mononuclear cells. Patient 1 is a compound heterozygote for the novel substitution c.256-1G > T that causes exon 2 skipping, leading to a premature stop codon in exon 3, and for the previously-known pathogenic mutation c.1183C > T (p.Arg395Cys). Patient 2 is homozygous for the novel mutation c.1185-1G > A that causes exon 7 skipping and the generation of a premature stop codon in exon 8, leading to the loss of the crucial adrenoxin binding domain of CYP27A1

Pichia pastoris regulates its gene-specific response to different carbon sources at the transcriptional, rather than the translational, level

Background: The methylotrophic, Crabtree-negative yeast Pichia pastoris is widely used as a heterologous protein production host. Strong inducible promoters derived from methanol utilization genes or constitutive glycolytic promoters are typically used to drive gene expression. Notably, genes involved in methanol utilization are not only repressed by the presence of glucose, but also by glycerol. This unusual regulatory behavior prompted us to study the regulation of carbon substrate utilization in different bioprocess conditions on a genome wide scale. Results: We performed microarray analysis on the total mRNA population as well as mRNA that had been fractionated according to ribosome occupancy. Translationally quiescent mRNAs were defined as being associated with single ribosomes (monosomes) and highly-translated mRNAs with multiple ribosomes (polysomes). We found that despite their lower growth rates, global translation was most active in methanol-grown P. pastoris cells, followed by excess glycerol- or glucose-grown cells. Transcript-specific translational responses were found to be minimal, while extensive transcriptional regulation was observed for cells grown on different carbon sources. Due to their respiratory metabolism, cells grown in excess glucose or glycerol had very similar expression profiles. Genes subject to glucose repression were mainly involved in the metabolism of alternative carbon sources including the control of glycerol uptake and metabolism. Peroxisomal and methanol utilization genes were confirmed to be subject to carbon substrate repression in excess glucose or glycerol, but were found to be strongly de-repressed in limiting glucose-conditions (as are often applied in fed batch cultivations) in addition to induction by methanol. Conclusions: P. pastoris cells grown in excess glycerol or glucose have similar transcript profiles in contrast to S. cerevisiae cells, in which the transcriptional response to these carbon sources is very different. The main response to different growth conditions in P. pastoris is transcriptional; translational regulation was not transcript-specific. The high proportion of mRNAs associated with polysomes in methanol-grown cells is a major finding of this study; it reveals that high productivity during methanol induction is directly linked to the growth condition and not only to promoter strength

Welfare status of cultured seabass (Dicentrarchus labrax L.) and seabream (Sparus aurata L.), assessed by blood parameters and tissue characteristics.

Seabass (SBS) and seabream (SBM) juveniles were reared with the goal of obtaining three different final densities (SINT 40 kg m3; INT 20 kg m3; SEM (0.2 kg m3) to ascertain the effects thereof on the welfare of the fish. Significant blood metabolites and hepatic glycogen were determined every 3 months and at harvest. Trials lasted 18 months in seabass and 17 months in seabream. At the end of experiment the main biometric productive parameters and quality of body composition were also recorded. Regarding intensively reared seabass (SINT-SBS, INT-SBS), the plasma triglycerids, total cholesterol and transaminases (AST, ALT) were always significantly higher than in semi-intensively maintained fish (SEM-SBS). At the final sampling in the SINT-SBS batch, the total protein and glucose were also markedly increased. Conversely, at harvest the liver glycogen content decreased in SINT-SBS (34±8 mg g liver) with respect to INT-SBS (57 ± 12 mg g)1 liver) and SEM-SBS (63±11 mg g liver). No differences among groups were observed for creatine kinase (CK) and lactate dehydrogenase (LDH). With regard to seabream, SINT-SBM and INT-SBM constantly showed plasma triglycerids and total cholesterol as being significantly higher when compared to SEM-SBM. In the final two blood samplings, SINT-SBM exhibited the most elevated values for LDH. At harvest, AST, ALT, total protein and glucose markedly increased in SINT-SBM, whereas liver glycogen content was reduced (22.5±9 mg g liver), more than in INT-SBM (70±16 mg g liver) and SEM-SBM (75±20 mg g liver). In both seabass and seabream, body composition was very similar in the different stocking densities, except for total cholesterol. Total n-3 polyunsaturated fatty acid (PUFA) in seabream was significantly different from fish of the semi-intensive groups; however, nutritional values and fatty acid profiles were equally good. The intermediate final density of seabass and seabream at 20 kg m3 seemed to give the best results in terms of their well being when compared to fish reared at 40 kg m3. The absence of differences in blood metabolites and hepatic glycogen levels between the intensive batch and the semi-intensive groups until harvest was a reference to the positive status of the fish. A density of 20 kg m3 can be considered acceptable for farm strategy planning