Screening and exclusion of Zika virus infection in travellers by an NS1-based ELISA and qRT-PCR.

OBJECTIVES: Zika virus (ZIKV) infection during pregnancy may cause neurological abnormalities in the foetus, and therefore fast and accurate laboratory assays are critical for rapid diagnosis. ELISA based on ZIKV NS1 protein has been developed and shown to be sensitive and highly specific; however, its negative and positive predictive values have not been tested. In this study we evaluated the ability of the NS1-based ELISA to exclude ZIKV infection and serve as a first-line screening tool for travellers. METHODS: We tested samples obtained during the peak of ZIKV infection from 1188 symptomatic and asymptomatic Israeli travellers using NS1-based IgG and IgM ELISA, real-time RT-PCR analysis and ZIKV neutralization. The Kaplan-Maier method was used to evaluate the duration of ZIKV RNA in whole blood and urine samples. RESULTS: NS1-based ELISA identified 20 true-positive, five false-positive and four false-negative cases, resulting in sensitivity and specificity of 83.3% (95%CI: 62-94%) and 97.5% (95%CI: 94-99%) respectively, and positive and negative predictive values of 80% (95%CI: 59-92%) and 98% (95%CI: 95-99%) respectively. Based on 14 RT-PCR-positive cases, median time to detect ZIKV RNA in whole blood was 17.5 days (range 5-58 days) and in urine 10 days (range 5-26 days). CONCLUSIONS: The NS1-based ELISA and RT-PCR in whole blood are highly reliable for identification of ZIKV-negative and -positive cases, respectively. Combination of both assays minimizes the risk of false-negative results, and thus allows the exclusion of ZIKV infection in travellers returning from ZIKV-endemic countries, including those who are pregnant or wish for preconception screening.

Sodium phosphate enemas do not worsen renal function among hospitalized patients with mild to moderate renal failure: a matched, case-control study.

BACKGROUND: Sodium phosphate enemas (SPEs) are widely used among hospitalized patients despite their potential to worsen renal failure. AIM: We decided to assess the extent to which this side effect is clinically relevant. DESIGN: We conducted a matched case-control, retrospective study in a cohort of hospitalized patients. METHODS: Patients treated and untreated with SPEs were matched for age, gender, baseline creatinine, usage of certain medications and several background diagnoses. Three groups of matched patients (whole study cohort, patients with baseline creatinine > 1.5 mg/dl and those with baseline creatinine > 2 mg/dl) were compared with regards to their creatinine and blood electrolyte concentrations during 3 consecutive hospitalization days after SPE application. RESULTS: Four hundred and twelve patients were included in this study of which 206 were treated by single SPEs. Exact matching was done for the whole study cohort, for 108 patients with baseline creatinine > 1.5 mg/dl and for 58 patients with baseline creatinine > 2 mg/dl. During 3 consecutive days after SPEs, the maximal blood concentrations of creatinine, phosphor and potassium did not differ significantly between treated patients and matched controls, in all three patients' groups. CONCLUSION: Application of SPEs neither seem to worsen mild to moderate renal failure, nor are associated with hyperphosphatemia or hyperkalemia in patients hospitalized in internal medicine departments.

Clinical outcomes and treatment approach for community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections in Israel.

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections are increasingly documented worldwide. We recently identified two major CA-MRSA clones in Israel: USA300 and t991. Here, we assessed clinical outcomes by CA-MRSA clones and the physicians' treatment approach to CA-MRSA infections. All community-onset, clinical MRSA isolates detected during 2011-2013 by Maccabi Healthcare Services were collected and characterized phenotypically and genotypically; data were collected retrospectively from electronic medical records. Of 309 patients with MRSA infections, 64 were identified as CA-MRSA (21 %). Of the CA-MRSA infections, 72 % had skin and soft tissue infections (SSTIs), 38 % were Panton-Valentine leukocidin (PVL)+, the major clone being USA300 (n = 13, 54 %). Of PVL- isolates (n = 40, 62 %), t991 was the major clone. Age was the only predictor for PVL+ CA-MRSA infection (p < 0.001). Patients with PVL+ CA-MRSA had higher incidence of SSTI recurrences (1.061 vs. 0.647 events per patient/per year, p < 0.0001) and were more likely to have the SSTI drained (64 % vs. 21 %, p = 0.003) when compared to PVL- CA-MRSA. USA300 was more common among adults, while t991 was more common among children (p = 0.002). The physician's referral to culture results and susceptibility were the only predictors of appropriate antibiotic therapy (p < 0.001). However, only a minority of physicians referred to culture results, regardless of subspecialties. PVL+ CA-MRSA isolates caused significantly more recurrences of SSTIs and increased the need for drainage compared with PVL- isolates. Physicians' awareness of CA-MRSA as a cause of SSTIs in the community was suboptimal. Culturing of pus-producing SSTIs is crucial for providing adequate antimicrobials and elucidating MRSA epidemiology.

Molecular epidemiology of community-onset methicillin-resistant Staphylococcus aureus infections in Israel.

Data on community-associated (CA) methicillin-resistant Staphylococcus aureus (MRSA) in Israel are scarce. The objective of this study was to characterize the major CA-MRSA clones in Israel. All clinical MRSA isolates detected in the community during a period of 2.5 years (2011-2013) from individuals insured by a major health maintenance organization in Israel were collected, with additional data from medical records. Antibiotic susceptibility patterns and staphylococcal chromosomal cassette mec (SCCmec) typing were determined. SCCmec IV and V isolates were further typed by pulsed-field gel electrophoresis (PFGE), spa typing, and detection of a panel of toxin genes. MRSA were detected in 280 patients, mostly from skin infections. Patients with SCCmec IV (n = 120, 43 %) were younger (p < 0.0001) and reported less contact with healthcare facilities. Almost all isolates were trimethoprim-sulfamethoxazole susceptible (98 %). spa-CC032, a typical nosocomial MRSA clone, accounted for 28 % of SCCmec IV. The two major CA-MRSA clones were t008 USA300 (13 %) and t991 (10 %); t991 was isolated mainly from children (75 %), was Panton-Valentine leukocidin (PVL) negative but eta-positive, and was typically susceptible to most antibiotic groups. PVL-positive strains (n = 31) included mainly USA300 (52 %) and t019 (13 %). While multiple genetic lineages were evident among community-onset MRSA in Israel, approximately 20 % are typical CA-MRSA clones, mainly USA300 and a local clone, t991.

Analytical characterisation of homoeopathic mother tinctures.

Quality of homoeopathic mother tinctures is assured by the definition of the starting material, the manufacturing process and the analytical characteristics described in the monograph. Traditionally analytical characterisation of the mother tincture comprises appearance, odour, identity, density and dry residue. According to annex I of directive 2001/83/EC an assay is only performed in case of a health hazard due to toxic compounds. The concept of marker substances as usually used in phytotherapy cannot be transferred to mother tinctures without research effort. For example the marker substances echinacoside, apigenin-7-glucoside and rosmarinic acid found in dried underground parts of Echinacea pallida Nutt., dried flower heads of Matricaria recutita L. and dried herb of Pulmonaria officinalis L. cannot be found in homoeopathic mother tinctures prepared from fresh material thereof.

Treatment of rats with the Pelargonium sidoides extract EPs 7630 has no effect on blood coagulation parameters or on the pharmacokinetics of warfarin.

Umckaloabo is a herbal drug for the treatment of respiratory tract infections. It contains an aqueous ethanolic extract from roots of Pelargonium sidoides DC (EPs) 7630) as the active ingredient. Polymeric polyphenols and coumarins have been identified as the principal ingredients of EPs 7630. In view of the coumarin content, it has been suggested that the administration of Umckaloabo could possibly be associated with an increased risk of bleeding. This study, therefore, investigated whether a change in blood coagulation parameters or an interaction with coumarin-type anticoagulants occurred after administration of EPs 7630 to rats. No effect on thromboplastin time (TPT), partial TPT (PTPT) or thrombin time (TT) was observed after oral administration of EPs 7630 (10, 75, 500 mg/kg) for 2 weeks, while treatment with warfarin (0.05 mg/kg) for the same period resulted in significant changes in TPT and PTPT. If EPs 7630 (500 mg/kg) and warfarin (0.05 mg/kg) were given concomitantly, the anticoagulant action of warfarin was not influenced. Similarly, the pharmacokinetics of warfarin were unchanged after pre-treatment with EPs 7630 for 2 weeks. Coumarin-type anticoagulants inhibit the synthesis of vitamin K-dependent coagulation factors via an identical mechanism in rat and man, and have a similar pattern of metabolism in both species. Moreover, as the coumarins so far identified in EPs 7630 do not posses the structural characteristics needed for anticoagulant activity, it appears unlikely that an increased tendency to haemorrhage arises in patients after intake of Umckaloabo.

Quality criteria of homoeopathic mother tinctures: considerations regarding suitable tests for homoeopathic monographs.

Quantitative determination of markers may improve quality control of herbal homoeopathic mother tinctures. Since the activity of homoeopathic medicinal products does not depend on the specific content of such markers, appropriate substances can be selected according to analytical aspects only. We tested carbohydrates, amino acids, total polyphenols and flavonols in different mother tinctures. The results obtained with the latter two groups of substances imply that corresponding tests might be useful for inclusion into pharmacopoeial monographs.

Pharmacokinetics of Ginkgo biloba extracts.

EGb 761 (definition see editorial) Ginkgo extract is produced by a validated production process. Its pharmacologically active constituents, flavonol glycosides and terpene lactones, are kept within a narrow range of 22 to 27 % and 5 to 7 %, respectively, by standardisation. The concentration of ginkgolic acids is below 5 ppm. The constant production process also maintains the concentrations of other constituents such as proanthocyanidins, carboxylic acids and non-flavone glycosides at a fairly constant level. In this article, we will summarise the data on the pharmacokinetics of flavonol glycosides and terpene lactones.

Influence of pharmaceutical quality on the bioavailability of active components from Ginkgo biloba preparations.

To be effective, herbal medicinal products are expected to meet comparable standards concerning the assessment of efficacy, safety and biopharmaceutical quality as chemically defined synthetic drugs as food supplements. However, these requirements are often not fulfilled, particularly regarding the characterization of biopharmaceutical properties such as in-vitro dissolution and in-vivo bioavailability. With respect to the relevance of biopharmaceutical quality of herbal medicinal products, two different Ginkgo biloba brands (test product: Ginkgo biloba capsules; reference product: Ginkgold) were analysed for dissolution rates and bioavailability of the most relevant active ingredients. Dissolution rates at pH 1 and 4.5 were determined according to the USP 23. The relative bioavailability of ginkgolide A, ginkgolide B and bilobalide was investigated after single oral administration of 120 mg Ginkgo biloba extract as tablets or capsules. Bioavailability data (area under the curve and peak concentration in plasma) were clearly different and did not show bioequivalence of test and reference products. The slow in-vitro dissolution of the test product resulted in a large decrease in bioavailability. These results indicate for the first time that the pharmaceutical properties of a herbal medicinal product have a significant impact on the rate and extent of drug absorption, and very likely on efficacy in humans.

Stimulation of glutamate, aspartate and gamma-aminobutyric acid release from synaptosomes by hyperforin.

Hyperforin is one pharmacologically active constituent of the medicinal herb Hypericum perforatum. The mechanism of its antidepressant-like activity is currently considered to be the inhibition of synaptic reuptake of neurotransmitters. Here, we will demonstrate that it also stimulates the release of glutamate from rat cortical synaptosomes, and that this effect is preceded by an increase in their free calcium [Ca2+]i levels. These hyperforin-related effects were also observed in the absence of Ca2+ in the medium. Although we noted enhanced glutamate, aspartate and GABA release under the influence of hyperforin, the release of various other amino acids was not enhanced. In contrast, reserpine did not influence the release of any of the amino acids studied. Adding hyperforin to synaptosomal suspension decreased their pHi, which returned to basal levels under certain incubation conditions. It also prevented the generation of ATP-induced pH gradients of isolated synaptic vesicles, and preformed pH-gradients were reversed by it. We will discuss the implications of our studies in understanding the mechanisms of hyperforin activity in relation to current findings on its pharmacological activity profile. Our observations suggest that neurotransmitter release stimulation from synaptosomes and the previously reported reuptake inhibitory properties of hyperforin are consequences of its effects on synaptosomal ionic homeostasis, and that it is not a reserpine-like agent.

Bioavailability of ginkgolides and bilobalide from extracts of ginkgo biloba using GC/MS.

The bioavailability of ginkgolides A, B and bilobalide was studied in rats after single oral administration of 30, 55 and 100 mg/kg Ginkgo extract EGb 761. The plasma levels of the terpene lactones were measured by a specific GC/MS method. The pharmacokinetics of the mentioned substances were found to be dose-linear. For the lowest dose maximum concentrations were 68, 40 and 159 ng/ml and half-lives 1.7, 2.0 and 2.2 h for ginkgolide A, B and bilobalide, respectively. Clearance values ranged from 24.2 to 37.6 ml/min/kg.

Oral bioavailability of hyperforin from hypericum extracts in rats and human volunteers.

Validated analytical methods suitable for determining hyperforin in plasma after administration of alcoholic Hypericum perforatum extracts containing hyperforin are described. After oral administration of 300 mg/kg Hypericum extract (WS 5572, containing 5% hyperforin) to rats maximum plasma levels of approximately 370 ng/ml (approx. 690 nM) were reached after 3 h, as quantified by a HPLC and UV detection method. Estimated half-life and clearance values were 6 h and 70 ml/min/kg respectively. Since therapeutic doses of Hypericum extracts are much lower than that used in rats, a more sensitive LC/MS/MS method was developed. The lower limit of quantification of this method was 1 ng/ml. Using this method, plasma levels of hyperforin could be followed for up to 24 h in healthy volunteers after administration of film coated tablets containing 300 mg hypericum extracts representing 14.8 mg hyperforin. The maximum plasma levels of approximately 150 ng/ml (approx. 280 nM) were reached 3.5 h after administration. Half-life and mean residence time were 9 and 12 h respectively. Hyperforin pharmacokinetics were linear up to 600 mg of the extract. Increasing the doses to 900 or 1200 mg of extract resulted in lower Cmax and AUC values than those expected from linear extrapolation of data from lower doses. Plasma concentration curves in volunteers fitted well in an open two-compartment model. In a repeated dose study, no accumulation of hyperforin in plasma was observed. Using the observed AUC values from the repeated dose study, the estimated steady state plasma concentrations of hyperforin after 3 x 300 mg/day of the extract, i.e., after normal therapeutic dose regimen, was approximately 100 ng/ml (approx. 180 nM).

Pharmacokinetics of [14C]-labelled Losigamone and enantiomers after oral administration to healthy subjects.

Losigamone ((+/-)-(R*,S*)-5-(2-chlorophenylhydroxymethyl)-4-methoxy-2 (5H)-furanone; AO-33) is a new potential antiepileptic drug undergoing clinical development. In a crossover study, 200 mg [14C]-labelled Losigamone, as well as 100 mg of each of the unlabelled enantiomers, was administered to 5 healthy volunteers as an oral suspension. The objectives of the study were to determine the mode of elimination, the excretion balance, metabolic profile, the in vitro and in vivo binding to plasma proteins and the pharmacokinetics of both enantiomers in plasma. From the plasma concentration-time profiles of [14C]-radioactivity and unchanged Losigamone it can be concluded that the absorption of Losigamone occurs very rapidly and the plasma concentration of the parent compound versus total radioactivity was consistently about 40%. An overall recovery of total radioactivity of about 97% with 85% in urine and 12% in faeces was found. Protein binding was 50%. Losigamone was extensively metabolized, with only traces of unchanged drug found in urine. The predominant metabolic pathways are hydroxylation and conjugation. After administration of the pure enantiomers, significant differences in pharmacokinetics were observed. The mean oral clearance of the (-)-enantiomer was 1863 ml/min and of the (+)-enantiomer was 171 ml/min. There was no chiral inversion after administration of the enantiomers.

KA 672-HCl, a neuronal activator against dementia: tolerability, safety, and preliminary pharmacokinetics after single and multiple oral doses in healthy male and female volunteers.

In vitro and animal experiments have characterized KA 672-HCl as a potent functional antagonist of excitatory amino acid-induced convulsions and mortality. In receptor-binding studies, the compound displayed high affinities to several serotoninergic, adrenergic, and dopaminergic receptors and to the sigma receptor. The potential for short- and long-term toxicity of KA 672-HCl in rats and dogs was found to be low. Double-blind, randomized, placebo-controlled studies were undertaken in healthy volunteers ranging from 52 to 74 years of age to determine tolerability, safety, and preliminary pharmacokinetics of single and repeated doses in humans. Single doses up to 40 mg were well tolerated, with no difference in effect from placebo. At 60 mg, approximately half of the volunteers experienced a moderate drug-related orthostatic syndrome. After repeated doses of 10 or 20 mg KA 672-HCl for 14 days only minor adverse events of mild intensity were reported with no clear relation to dose or a clinically relevant difference from placebo. A mild decrease in semisupine and standing blood pressure 4 hours after administration was observed in the 20 mg group with no occurrence of orthostasis. Linear pharmacokinetics were observed after repeated doses. However, this was not the case after single-dose administration, as generally higher plasma concentrations were observed after the 20-mg dose than would have been predicted from the 10-mg data. The mean terminal phase half-life after the 20 mg dose was 11.1 hours and 13.7 hours after repeated and single doses, respectively. The safety and tolerability data support a continuation of therapeutic trials. KA 672-HCl is currently entering phase II development.

[Pharmaokinetics of beta-escin after administration of various Aesculus extract containing formulations]. / Pharmakokinetik von beta-Aescin nach Gabe verschiedener Aesculus-Extrakt enthaltender Formulierungen.

With a specific radioimmunoassay the pharmacokinetics and relative bioavailability of escin was measured after administration of different formulations containing Aesculus-extract. Of special interest was the relative bioavailability of escin after administration of a newly developed film-coated tablet with sustained release in comparison to a reference formulation. In a cross-over steady-state study in 24 volunteers bioequivalence of test and reference preparation could be demonstrated. The 90% confidence interval of the AUC (O-tau) was 98.3 to 120.9%.

Antiviral and antiphlogistic activities of Hamamelis virginiana bark.

A crude hydroalcoholic extract from Hamamelis virginiana bark was subjected to ultrafiltration (UF) with a cut-off limit of 3 kDa to obtain a higher and a lower molecular weight fraction. Characterisation of the fractions was attempted with TLC, HPLC, acidic hydrolysis, and chromatography over Sephadex LH-20. The UF-concentrate was shown to consist mainly of oligomeric to polymeric proanthocyanidins (PA). This fraction was found to exhibit significant antiviral activity against Herpes simplex virus type 1 (HSV-1). In addition, the UV-concentrate displayed radical scavenging properties, inhibited alpha-glucosidase as well as human leukocyte elastase (HLE), and exhibited strong antiphlogistic effects in the croton oil ear edema test in the mouse. With the exception of the antioxidant potential and the inhibition of HLE-action the lower molecular fraction possessed weaker activities and contained mainly hamamelitannin, catechin, and further, unidentified constituents.

Enzymatic cleavage of thymopoietin oligopeptides by pancreatic and intestinal brush-border enzymes.

The intestinal enzymatic degradation of the immunomodulating peptides thymotrinan (TP3), thymocartin (TP4), and thymopentin (TP5), three oligopeptides derived from the naturally occurring thymus hormone thymopoietin, was investigated to evaluate their potential for peroral drug delivery. In the presence of brush-border membrane vesicles, crude pancreas extract and everted rings from duodenum, jejunum, ileum, and colon, all peptides were shown to be degraded both by pancreatic enzymes and brush-border aminopeptidases. Degradation clearances (Cldeg) of TP3, TP4, and TP5 were calculated for a quantitative comparison of peptide stability. In the presence of crude pancreas extract, there was a rapid degradation of TP5 (Cldeg 17.9 ml/min) in comparison with TP3 and TP4 (Cldeg 0.95 and 0.56 ml/min, respectively, at 0.2 mM peptide concentration) caused by the cleavage of the C-terminal tyrosine by carboxypeptidase A, whereas TP3 and TP4 underwent hydrolysis by aminopeptidase N. In the presence of brush-border membrane vesicles, the degradation clearances were 3.9, 3.1, and 2.4 ml/min at 0.2 mM concentrations of TP4, TP5, and TP3, respectively. The clearance of all peptides was lowered with increasing peptide concentrations, indicating saturable degradation processes. The degradation of the thymopoietin oligopeptides in the presence of brush-border membrane enzymes was exclusively catalyzed by aminopeptidase N. The degradation of all peptides was highly dependent on the intestinal segment, with the lowest degradation clearance observed in the colon.

Pharmacokinetics of losigamone, a new antiepileptic drug, in healthy male volunteers.

Losigamone (AO-33), a new potential antiepileptic drug, was tested in 52 healthy male volunteers in 4 placebo-controlled phase I studies. In study 1 single doses of 100, 200, 300, 500, 700, and 1,000 mg losigamone were given as a fast releasing capsule to 12 subjects. The pharmacokinetics of losigamone measured after administration of 100, 300, and 700 mg was linear. Clearance and t1/2 were about 350 ml/min and 4 h, respectively, the Cmax values of 0.7, 1.7, and 4.4 micrograms/ml were reached after 2.5 h. In study 2,500 mg losigamone were given as a fast release capsule for 6 days (t.i.d.) to 12 subjects. There was a small but statistically significant decrease for the AUC but no change in t1/2, Cmax or tmax comparing single dose kinetics on day 1 and 8. There appeared to be no change in caffeine clearance on days 1 and 9. Study 2 was repeated in 20 volunteers with a film-coated tablet. Pharmacokinetic parameters appeared to be unaffected by this change in galenical formulation. In study 4 daily doses of 400, 1,200, and 1,800 mg losigamone were given 28 days to 24 subjects. The kinetics of caffeine and antipyrine were compared on days 1 and 29. With the exception of t1/2 for antipyrine in the 400 mg group there was no statistically significant change in pharmacokinetic parameters. Generally, losigamone was well tolerated and no serious adverse side-effects occurred. In some subjects a reversible increase in transaminases was observed.

[Pharmacokinetic properties of Bilobalide and Ginkgolides A and B in healthy subjects after intravenous and oral administration of Ginkgo biloba extract (EGb 761)]. / Propriétés pharmacocinétiques du Bilobalide et des Ginkgolides A et B chez le sujet sain après administrations intraveineuses et orales d'extrait de Ginkgo biloba (EGb 761).

The pharmacokinetics of Ginkgolide A, Ginkgolide B and Bilobalide, which are compounds extracted from the dried leaves of the Ginkgo biloba tree, were investigated in 12 young healthy volunteers (six men and six women; mean +/- SD age = 25 +/- 5 years) after single-dose administration of Ginkgo biloba extract. Subjects were given, on three occasions, Ginkgo biloba extract as a solution either orally (in fasting conditions and after a standard meal) or intravenously; corresponding to single doses of Ginkgolide A, Ginkgolide B and Bilobalide ranging from 0.90 mg to 3.36 mg. After each dosing, blood and urine samples were collected for up to 36 h and 48 h, for measurements of Ginkgolide A, Ginkgolide B and Bilobalide. Plasma and urine concentrations of these compounds were quantitatively measured by gas chromatography/mass spectrometry using negative chemical ionization, by applying a very sensitive method which allowed plasma concentrations as low as 0.2 ng/ml of each compound to be measured. When given orally, while fasting, the extents of bioavailability are high, as shown by bioavailability coefficients (FAUC) mean (+/- SD) values equal to 0.80 (+/- 0.09), 0.88 (+/- 0.21) and 0.79 (+/- 0.30) for Ginkgolide A, Ginkgolide B and Bilobalide respectively. Food intake does not change AUC quantitatively but increases Tmax. For the three compounds of interest, after oral dosing while fasting, differences can be noted for the elimination half-lives (T1/2Z), which exhibit mean values equal to 4.50, 10.57 and 3.21 h, as well as mean residence times (MRT), equal to 5.86, 11.25 and 4.89 h, for Ginkgolide A, Ginkgolide B and Bilobalide respectively.(ABSTRACT TRUNCATED AT 250 WORDS)