The aim of the present study was to characterize, quantify, and compare the different selenium species that are produced when lactic fermentation with two different types of microorganisms, bacteria ...(Lactobacillus) and yeast (Saccharomyces), take place to produce yogurt and kefir, respectively, and to study the transformation process of these species as a function of time. These two dairy products were chosen for the study because they are highly consumed in different cultures. Moreover, the microorganisms present in the fermentation processes are different. While the bacteria Lactobacillus is the one responsible for yogurt fermentation, a partnership between bacteria and the yeast Saccharomyces causes kefir fermentation. A comparative study has been carried out by fermenting Se(IV) enriched milk in the presence of both types of microorganisms, where the concentration range studied was from 0.5 to 20 μg g-1. Enzymatic extraction enabled selenium speciation profiles, obtained by anionic exchange and ion-pairing reversed phase high performance liquid chromatography (IP-RP-HPLC) with inductively coupled plasma mass spectrometry (ICP-MS) detection. Scanning electron microscopy (SEM) applied to the enriched samples showed segregated Se0, at added concentrations higher than 5 μg g-1. The main Se species formed depended on the type of microorganism involved in the fermentation process, SeCys2 and MeSeCys being the main species generated in yogurt and SeMet in kefir. The results obtained are different for both kinds of samples. Lactic fermentation for yogurt produced an increment in selenocystine (SeCys2) and Se-methylselenocysteine (MeSeCys), while fermentation to produce kefir also incremented the selenomethionine (SeMet) concentration. The Se species are stable for at least 10 and 15 days for kefir and yogurt, respectively. After this period, selenocystine concentration decreased, and the concentration of Se-methylselenocysteine was found to significantly increase.
Summary
Background
Suboptimal response to ursodeoxycholic acid occurs in 40% of primary biliary cholangitis (PBC) patients, affecting survival. Achieving a deep response (normalisation of alkaline ...phosphatase ALP and bilirubin ≤0.6 upper limit of normal) improves survival. Yet, the long‐term effectiveness of second‐line treatments remains uncertain.
Aims
To evaluate the long‐term effectiveness of obeticholic acid (OCA) ± fibrates. Focusing on biochemical response (ALP ≤1.67 times the upper limit of normal, with a decrease of at least 15% from baseline and normal bilirubin levels), normalisation of ALP, deep response and biochemical remission (deep response plus aminotransferase normalisation).
Methods
We conducted a longitudinal, observational, multicentre study involving ursodeoxyccholic acid non‐responsive PBC patients (Paris‐II criteria) from Spain and Portugal who received OCA ± fibrates.
Results
Of 255 patients, median follow‐up was 35.1 months (IQR: 20.2–53). The biochemical response in the whole cohort was 47.2%, 61.4% and 68.6% at 12, 24 and 36 months. GLOBE‐PBC and 5‐year UK‐PBC scores improved (p < 0.001). Triple therapy (ursodeoxycholic acid plus OCA plus fibrates) had significantly higher response rates than dual therapy (p = 0.001), including ALP normalisation, deep response and biochemical remission (p < 0.001). In multivariate analysis, triple therapy remained independently associated with biochemical response (p = 0.024), alkaline phosphatase normalisation, deep response and biochemical remission (p < 0.001). Adverse effects occurred in 41.2% of cases, leading to 18.8% discontinuing OCA. Out of 55 patients with cirrhosis, 12 developed decompensation. All with baseline portal hypertension.
Conclusion
Triple therapy was superior in achieving therapeutic goals in UDCA‐nonresponsive PBC. Decompensation was linked to pre‐existing portal hypertension.
Longitudinal, real‐world study on 255 UDCA‐nonresponsive PBC patients (Per Paris II criteria); median follow‐up of 35.1 months (IQR: 20–53). All patients received obeticholic acid (OCA), with 25% receiving later add‐on fibrate treatment (triple therapy). In multivariate analysis, triple therapy outperformed dual therapy across all surrogate biochemical endpoints of outcomes.
The suitability of Se-enriched fermented milk as a food product for humans has been studied and sensory features, survival of microorganisms in the presence of selenium and Se bioaccessibility are ...presented. Sensory features and the number of colonies of microorganisms observed until the 4th week after fermentation were not affected at Se concentration levels below 2
μg
g
−1. To investigate bioaccessible selenium-containing compounds in selenized fermented milk, total Se was quantified after in vitro gastrointestinal digestion. Results showed that 76
±
3% Se was extracted after gastrointestinal digestion and 24
±
6% remained in the insoluble residue. To identify the main species of bioaccessible Se of low molecular weight (MW) generated during gastrointestinal digestion, liquid chromatography-inductively coupled plasma-mass spectrometry (LC-ICP-MS) was used. Results showed that identified selenocompounds were smaller than 1.5
kDa, with selenocystine and Se-methylselenocysteine being the most abundant compounds present in the extracts.
Selenium species present in water-soluble fractions of proteins of lyophilised biological samples, such as fish (tuna and trout), shellfish (krill, oyster and mussel), vegetal tissues (white clover ...and wheat flour) and selenized yeast were evaluated. The water-soluble fractions with and without 4% (w/v) sodium dodecyl sulphate were analysed by size exclusion chromatography coupled to both inductively coupled plasma–mass spectrometry (SEC–ICP–MS) and UV spectrophotometers. Two different columns with effective separation ranges between 300 and 1
kDa (Biosep-SEC-2000) and 7 and 0.1
kDa (Superdex Peptide HR 10/30) were used, with 5
mM sodium phosphate at pH 7.2 (0.8
ml
min
−1) and 5
mM sodium phosphate with 60
mM NaCl at pH 7.5 (0.6
ml
min
−1) as mobile phases, respectively.
The enzymatic digests of both the water-soluble fractions and the non-soluble residues were analysed by cation exchange chromatography–ICP–MS, using a Hamilton PRP-X200 column and 4
mM pyridine (1
ml
min
−1) at pH 2.8 and 4.7 as mobile phases.
The presence of high molecular weight selenoproteins (150–50
kDa) in the samples studied, except for fish and white clover with proteins <30
kDa, was demonstrated and low molecular weight (<5
kDa) compounds were also found.
Selenomethionine species (SeMet) within the range of 0.2–600
μg
g
−1 was quantified in the selected tissues, although selenocystine (SeCys
2) was identified only in vegetal samples (0.1–0.7
μg
g
−1). Trimethylselenonium ion was quantified (0.1–0.3
μg
g
−1) in oyster, mussel, trout and white clover tissues. Inorganic selenium as selenite was found in the selenized yeast, krill, and white clover samples (0.2–3
μg
g
−1), while selenate was found to be present only in yeast tissues (8
μg
g
−1).
This study aims to demonstrate the applicability and importance of zebrafish (Danio rerio) model to study acute and chronic inflammatory responses induced by different stimuli: carrageenan phlogogen ...(nonimmune); acute infection by bacteria (immune); foreign body reaction (chronic inflammation by round glass coverslip implantation); reaction induced by xenotransplantation. In addition to the advantages of presenting low breeding cost, high prolificity, transparent embryos, high number of individuals belonging to the same spawning and high genetic similarity that favor translational responses to vertebrate organisms like humans, zebrafish proved to be an excellent tool, allowing the evaluation of edema formation, accumulation of inflammatory cells in the exudate, mediators, signaling pathways, gene expression and production of specific proteins. Our studies demonstrated the versatility of fish models to investigate the inflammatory response and its pathophysiology, essential for the successful development of studies to discover innovative pharmacological strategies.
Despite the increasing evidence of the benefit of corticosteroids for the treatment of moderate-severe coronavirus disease 2019 (COVID-19) patients, no data are available about the potential role of ...high doses of steroids for these patients. We evaluated the mortality, the risk of need for mechanical ventilation (MV), or death and the risk of developing a severe acute respiratory distress syndrome (ARDS) between high (HD) and standard doses (SD) among patients with a severe COVID-19. All consecutive confirmed COVID-19 patients admitted to a single center were selected, including those treated with steroids and an ARDS. Patients were allocated to the HD (≥ 250 mg/day of methylprednisolone) of corticosteroids or the SD (≤ 1.5 mg/kg/day of methylprednisolone) at discretion of treating physician. Five hundred seventy-three patients were included: 428 (74.7%) men, with a median (IQR) age of 64 (54–73) years. In the HD group, a worse baseline respiratory situation was observed and male gender, older age, and comorbidities were significantly more common. After adjusting by baseline characteristics, HDs were associated with a higher mortality than SD (adjusted OR 2.46, 95% CI 1.59–3.81,
p
< 0.001) and with an increased risk of needing MV or death (adjusted OR 2.35,
p
= 0.001). Conversely, the risk of developing a severe ARDS was similar between groups. Interaction analysis showed that HD increased mortality exclusively in elderly patients. Our real-world experience advises against exceeding 1–1.5 mg/kg/day of corticosteroids for severe COVID-19 with an ARDS, especially in older subjects. This reinforces the rationale of modulating rather than suppressing immune responses in these patients.
Swine brucellosis by Brucella suis biovar 2 is an emerging disease whose control is based on serological testing and culling. However, current serological tests detect antibodies to the ...O-polysaccharide (O/PS) moiety of Brucella smooth lipopolysaccharide (S-LPS), and thus lack specificity when infections by Yersinia enterocolitica O:9 and other gram-negative bacteria carrying cross-reacting O/PS occur. The skin test with the protein-rich brucellin extract obtained from rough B. melitensis B115 is assumed to be specific for discriminating these false positive serological reactions (FPSR). However, B115 strain, although unable to synthesize S-LPS, accumulates O/PS internally, which could cause diagnostic problems. Since the brucellin skin test has been seldom used in pigs and FPSR are common in these animals, we assessed its performance using cytosoluble protein extracts obtained from B. abortus rough mutants in manBcore or per genes (critical for O/PS biosynthesis) and B. melitensis B115. The diagnostic sensitivity and specificity were determined in B. suis biovar 2 culture positive and brucellosis free sows, and apparent prevalence in sows of unknown individual bacteriological and serological status belonging to B. suis biovar 2 naturally infected herds. Moreover, the specificity in discriminating brucellosis from FPSR was assessed in brucellosis free boars showing FPSR. The skin test with B. abortus ΔmanBcore and B. melitensis B115 allergens performed similarly, and the former one resulted in 100% specificity when testing animals showing FPSR in indirect ELISA, Rose Bengal and complement fixation serological tests. We conclude that O/PS-free genetically defined mutants represent an appropriate alternative to obtain Brucella protein extracts for diagnosing swine brucellosis.
This paper reports a sensitive, highly selective flow injection (FI) method for the chemiluminescence-based determination of enrofloxacin (ENRO), a fluoroquinolone (FQ) antibiotic, in natural water ...samples. The detection principle is the on-line monitored photoreaction of tris(1,10-phenantroline)ruthenium(II) Ru(phen)32+ with peroxydisulphate ion to give Ru(phen)33+, which is then reduced by the antimicrobial producing the chemiluminescence (CL) signal. The low selectivity of the CL reaction required using solid-phase extraction with molecularly imprinted polymers (MISPE) in combination with the FI-CL system. Fluoroquinolone-selective MIP beads were obtained by acid etching of silica/MIP composites and packed into SPE cartridges to clean-up and preconcentrate samples prior injection into the FI-CL system. Various parameters influencing the CL signal intensity including pH, flow rate, and Ru(phen)32+ and peroxydisulphate concentrations, were optimized. The variables affecting the extraction efficiency of the polymers were also optimized in order to minimize non-specific interactions and enable the selective uptake of the antibiotics from real samples. CL measurements made over the pH range 3.8–4.2 allowed the selective determination of enrofloxacin, and measurements at pH4.8–5.8 the quantitation of total FQs (enrofloxacin, ciprofloxacin, norfloxacin, sarafloxacin, lomefloxacin and danofloxacin). The limit of detection for enrofloxacin was 0.27μgL−1 when 10mL water samples were preconcentrates. The proposed method was used to determine trace amounts of FQs in mineral and tap water, and also in water samples from the river Quibú (Cuba), all with good recoveries: 84–119% (RSD 2.1–9.3%, n=3) for samples fortified with enrofloxacin at concentrations from 4 to 40μgL−1. The method was validated by HPLC with fluorescence detection.
•A novel MISPE-FI-CL method for enrofloxacin analysis was developed using the Ru(phen)32+/S2O82− system.•Adequate selection of the carrier pH allows selective determination of enrofloxacin or fluoroquinolones screening•Other amine containing compounds present in natural water samples did not interfere.•The method was validated by HPLC-FLD and applied to tap and river water analysis.
Background and ImportanceThe pharmacokinetic/pharmacodynamic (PK/PD) target for vancomycin has recently been defined as an area under the curve (AUC) over 24 hours/minimum inhibitory concentration ...(MIC) of 400-600.Aim and ObjectivesTo evaluate the degree of concordance of recommendations after dose adjustment of vancomycin according to minimum plasma concentration (Cmin) and AUC/MIC ratio.Material and MethodsRetrospective study in adult patients who were treated with vancomycin administered by intermittent perfusion and monitored by the Pharmacy Service at a general hospital during the month of August 2022.Variables collected: sex, age, weight, height, glomerular filtration rate (according to Cockcroft-Gault), total daily dose and recommendation issued based on the determination of Cmin and AUC/MIC.Appropriate Cmin were considered 15-20µg/mL in complicated infection (endocarditis, nosocomial pneumonia, meningitis, osteomyelitis/osteoarticular infection and wound infection/abscess) and 10-15µg/mL in all other infections. For the calculation of AUC/MIC, MIC=1µg/mL was assumed. Interpretation of plasma level and individualised vancomycin adjustment was performed using MediWare Pharm++® software using a bicompartmental model and a single vancomycin level (Cmin).ResultsVancomycin treatment was initiated and monitored in 42 patients (52.4% female; 72.3 ± 12.3). Anthropometric parameters (weight: 81.8 ± 17.9kg; height: 163.8 ± 7.9cm; glomerular filtration rate: 61.5 ± 27.0ml/min/1.73m2); total daily dose: 1,878.5mg ± 524.8mg. The recommendation issued was concordant via Cmin and AUC/MIC in 35.7%. In the case of discordance, overexposure was observed in 66.6% of cases.Conclusion and RelevanceApproximately 2 out of 3 recommendations were discordant according to the method used, with a high number of overexposures observed in the case of recommendations based on Cmin. Therefore, despite the small sample size, the implementation of vancomycin therapeutic monitoring according to AUC is considered necessary for the optimisation of therapeutic management.References and/or AcknowledgementsConflict of InterestNo conflict of interest
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