Neutralizing Carbapenem Resistance by Co-Administering Meropenem with Novel ß-Lactam-Metallo-ß-Lactamase Inhibitors.

Virulent Enterobacterale strains expressing serine and metallo-ß-lactamases (MBL) genes have emerged responsible for conferring resistance to hard-to-treat infectious diseases. One strategy that exists is to develop ß-lactamase inhibitors to counter this resistance. Currently, serine ß-lactamase inhibitors (SBLIs) are in therapeutic use. However, an urgent global need for clinical metallo-ß-lactamase inhibitors (MBLIs) has become dire. To address this problem, this study evaluated BP2, a novel beta-lactam-derived ß-lactamase inhibitor, co-administered with meropenem. According to the antimicrobial susceptibility results, BP2 potentiates the synergistic activity of meropenem to a minimum inhibitory concentration (MIC) of ≤1 mg/L. In addition, BP2 is bactericidal over 24 h and safe to administer at the selected concentrations. Enzyme inhibition kinetics showed that BP2 had an apparent inhibitory constant (Kiapp) of 35.3 µM and 30.9 µM against New Delhi Metallo-ß-lactamase (NDM-1) and Verona Integron-encoded Metallo-ß-lactamase (VIM-2), respectively. BP2 did not interact with glyoxylase II enzyme up to 500 µM, indicating specific (MBL) binding. In a murine infection model, BP2 co-administered with meropenem was efficacious, observed by the >3 log10 reduction in K. pneumoniae NDM cfu/thigh. Given the promising pre-clinical results, BP2 is a suitable candidate for further research and development as an (MBLI).

In Vitro and In Vivo Development of a ß-Lactam-Metallo-ß-Lactamase Inhibitor: Targeting Carbapenem-Resistant Enterobacterales.

ß-lactams are the most prescribed class of antibiotics due to their potent, broad-spectrum antimicrobial activities. However, alarming rates of antimicrobial resistance now threaten the clinical relevance of these drugs, especially for the carbapenem-resistant Enterobacterales expressing metallo-ß-lactamases (MBLs). Antimicrobial agents that specifically target these enzymes to restore the efficacy of last resort ß-lactam drugs, that is, carbapenems, are therefore desperately needed. Herein, we present a cyclic zinc chelator covalently attached to a ß-lactam scaffold (cephalosporin), that is, BP1. Observations from in vitro assays (with seven MBL expressing bacteria from different geographies) have indicated that BP1 restored the efficacy of meropenem to ≤ 0.5 mg/L, with sterilizing activity occurring from 8 h postinoculation. Furthermore, BP1 was nontoxic against human hepatocarcinoma cells (IC50 > 1000 mg/L) and exhibited a potency of (Kiapp) 24.8 and 97.4 µM against Verona integron-encoded MBL (VIM-2) and New Delhi metallo ß-lactamase (NDM-1), respectively. There was no inhibition observed from BP1 with the human zinc-containing enzyme glyoxylase II up to 500 µM. Preliminary molecular docking of BP1 with NDM-1 and VIM-2 sheds light on BP1's mode of action. In Klebsiella pneumoniae NDM infected mice, BP1 coadministered with meropenem was efficacious in reducing the bacterial load by >3 log10 units' postinfection. The findings herein propose a favorable therapeutic combination strategy that restores the activity of the carbapenem antibiotic class and complements the few MBL inhibitors under development, with the ultimate goal of curbing antimicrobial resistance.

The in vitro and in vivo potential of metal-chelating agents as metallo-beta-lactamase inhibitors against carbapenem-resistant Enterobacterales.

The recent surge in beta-lactamase resistance has created superbugs, which pose a current and significant threat to public healthcare. This has created an urgent need to keep pace with the discovery of inhibitors that can inactivate these beta-lactamase producers. In this study, the in vitro and in vivo activity of 1,4,7-triazacyclononane-1,4,7 triacetic acid (NOTA)-a potential metallo-beta-lactamase (MBL) inhibitor was evaluated in combination with meropenem against MBL producing bacteria. Time-kill studies showed that NOTA restored the efficacy of meropenem against all bacterial strains tested. A murine infection model was then used to study the in vivo pharmacokinetics and efficacy of this metal chelator. The coadministration of NOTA and meropenem (100 mg/kg.bw each) resulted in a significant decrease in the colony-forming units of Klebsiella pneumoniae NDM-1 over an 8-h treatment period (>3 log10 units). The findings suggest that chelators, such as NOTA, hold strong potential for use as a MBL inhibitor in treating carbapenem-resistant Enterobacterale infections.

Mating success of timed pregnancies in Sprague Dawley rats: Considerations for execution.

This study compares three different mating techniques in Sprague-Dawley rats, using the pregnancy rate as the main indicator of success. It provides recommendations for timed-pregnancy experiments to achieve an appropriate sample size for the study of human pregnancy disorders. The implementation of a preconditioning phase, determination of the estrous cycle, the use of two mating strategies (Lee-Boot and Whitten effect), female: male mating ratios, and cohabitation duration should be considered as they improve the mating success rate.

Physiological characterization of an arginine vasopressin rat model of preeclampsia.

Rodent models have contributed greatly to our understanding of preeclampsia (PE) progression in humans, however to-date no model has been able to effectively replicate the clinical presentation of the disease. This study aimed to provide a thorough physiological characterization of the arginine vasopressin (AVP)-induced rat model of PE to determine its applicability in studying the pathophysiology of PE. Female Sprague Dawley rats (n = 24) were separated into four groups (n = 6 per group) viz., pregnant AVP, pregnant saline, non-pregnant AVP, and non-pregnant saline. All animals received a continuous dose of either AVP (150 ng/h) or saline via subcutaneous mini osmotic pumps for 18 days. Full physiological characterization of the model included measuring systolic and diastolic blood pressure, and collecting urine and blood samples for biochemical analysis. AVP infusion significantly increased blood pressure and urinary protein levels in the pregnant rats (p < 0.05). Biochemical markers measured, differed significantly in the AVP-treated vs the pregnant saline groups (p < 0.05). Placental and individual pup weight decreased significantly in the pregnant AVP vs pregnant saline group (p < 0.05). The physiological and hematological data confirm the usefulness of this rat model in the study of PE, since AVP-induced vasoconstriction increases peripheral resistance and successfully mimics the pathological changes associated with PE development in humans.Abbreviations: PE: preeclampsia; AVP: arginine vasopressin; ISSHP: International Society for the Study of Hypertension in Pregnancy; ACOG: American College of Obstetricians and Gynecologists; RUPP: reduced uterine perfusion pressure; sFlt-1: soluble fms-like tyrosine kinase; VEGF: vascular endothelial growth factor; PlGF: placental growth factor; AVP: arginine vasopressin; PAVP: pregnant AVP-treated; PS: pregnant saline; GD: gestational day; ALT: alanine transaminase; NAVP: non-pregnant AVP-treated; NS: non-pregnant saline; AST: aspartate aminotransferase; HDL: high-density lipoprotein; RBC: red blood cell; RAAS: renin-angiotensin aldosterone system; HELLP: hemolysis, elevated liver enzymes, low platelet.

Chitosan-Based Hydrogel for the Dual Delivery of Antimicrobial Agents Against Bacterial Methicillin-Resistant Staphylococcus aureus Biofilm-Infected Wounds.

Chronic wound infections caused by antibiotic-resistant bacteria have become a global health concern. This is attributed to the biofilm-forming ability of bacteria on wound surfaces, thus enabling their persistent growth. In most cases, it leads to morbidity and in severe cases mortality. Current conventional approaches used in the treatment of biofilm wounds are proving to be ineffective due to limitations such as the inability to penetrate the biofilm matrix; hence, biofilm-related wounds remain a challenge. Therefore, there is a need for more efficient alternate therapeutic interventions. Hydrogen peroxide (HP) is a known antibacterial/antibiofilm agent; however, prolonged delivery has been challenging due to its short half-life. In this study, we developed a hydrogel for the codelivery of HP and antimicrobial peptides (Ps) against bacteria, biofilms, and wound infection associated with biofilms. The hydrogel was prepared via the Michael addition technique, and the physiochemical properties were characterized. The safety, in vitro, and in vivo antibacterial/antibiofilm activity of the hydrogel was also investigated. Results showed that the hydrogel is biosafe. A greater antibacterial effect was observed with HP-loaded hydrogels (CS-HP; hydrogel loaded with HP and CS-HP-P; hydrogel loaded with HP and peptide) when compared to HP as seen in an approximately twofold and threefold decrease in minimum inhibitory concentration values against methicillin-resistant Staphylococcus aureus (MRSA) bacteria, respectively. Similarly, both the HP-releasing hydrogels showed enhanced antibiofilm activity in the in vivo study in mice models as seen in greater wound closure and enhanced wound healing in histomorphological analysis. Interestingly, the results revealed a synergistic antibacterial/antibiofilm effect between HP and P in both in vitro and in vivo studies. The successfully prepared HP-releasing hydrogels showed the potential to combat bacterial biofilm-related infections and enhance wound healing in mice models. These results suggest that the HP-releasing hydrogels may be a superior platform for eliminating bacterial biofilms without using antibiotics in the treatment of chronic MRSA wound infections, thus improving the quality of human health.

Alterations in neurotransmitter levels and transcription factor expression following intranasal buprenorphine administration.

Buprenorphine is an opioid drug used in the management of pain and the treatment opioid addiction. Like other opioids, it is believed that it achieves these effects by altering functional neurotransmitter pathways and the expression of important transcription factors; cyclic AMP response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF) in the brain. However, there is a lack of scientific evidence to support these theories. This study investigated the pharmacodynamic effects of BUP administration by assessing neurotransmitter and molecular changes in the healthy rodent brain. Sprague-Dawley rats (150-200 g) were intranasally administered buprenorphine (0.3 mg/mL) and sacrificed at different time points: 0.25, 0.5, 1, 2, 4, 6, 8 and 24 h post drug administration. LC-MS was used to quantify BUP and neurotransmitters (GABA, GLUT, DA, NE and 5-HT) in the brain, while CREB and BDNF gene expression was determined using qPCR. Results showed that BUP reached a Cmax of 1.21 ± 0.0523 ng/mL after 2 h, with all neurotransmitters showing an increase in their concentration over time, with GABA, GLUT and NE reaching their maximum concentration after 8 h. DA and 5-HT reached their maximum concentrations at 1 h and 24 h, respectively post drug administration. Treatment with BUP resulted in significant upregulation in BDNF expression throughout the treatment period while CREB showed patterns of significant upregulation at 2 and 8 h, and downregulation at 1 and 6 h. This study contributes to the understanding of the pharmacodynamic effects of BUP in opioid addiction by proving that the drug significantly influences NT pathways that are implicated in opioid addiction.

Mass Spectrometric Imaging of the Brain Demonstrates the Regional Displacement of 6-Monoacetylmorphine by Naloxone.

Overdose is the main cause of mortality among heroin users. Many of these overdose-induced deaths can be prevented through the timely administration of naloxone (NLX), a nonselective mu (µ)-, kappa (κ)-, and delta (δ)-opioid receptor antagonist. NLX competitively inhibits opioid-overdose-induced respiratory depression without eliciting any narcotic effect itself. The aim of this study was to investigate the antagonistic action of NLX by comparing its distribution to that of 6-monacetylmorphine (6-MAM), heroin's major metabolite, in a rodent model using mass spectrometric imaging (MSI) in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Male Sprague-Dawley rats (n = 5) received heroin (10 mg kg-1) intraperitoneally, NLX (10 mg kg-1) intranasally, and NLX injected intranasally 5 min after heroin administration. The animals were sacrificed 15 min after dose and brain tissues were harvested. The MSI image analysis showed a region-specific distribution of 6-MAM in the brain regions including the corpus callosum, hippocampal formation, cerebral cortex, corticospinal tracts, caudate putamen, thalamus, globus pallidus, hypothalamus, and basal forebrain regions of the brain. The antagonist had a similar biodistribution throughout the brain in both groups of animals that received NLX or NLX after heroin administration. The MSI analysis demonstrated that the intensity of 6-MAM in these brain regions was reduced following NLX treatment. The decrease in 6-MAM intensity was caused by its displacement by the antagonist and its binding to these receptors in these specific brain regions, consequently enhancing the opioid elimination. These findings will contribute to the evaluation of other narcotic antagonists that might be considered for use in the treatment of drug overdose via MSI.

Fusaric acid alters Akt and ampk signalling in c57bl/6 mice brain tissue.

The brain is a highly metabolic organ and requires regulatory mechanisms to meet its high energy demand, with the PI3K/Akt and AMPK signalling pathways being central regulators of cellular energy and metabolism, also making them major targets for the development of neurometabolic disorders. Fusaric acid (FA), a toxin of fungal origin, was found to be a potent hypotensive agent in vivo and in clinical trials by altering brain neurochemistry thus demonstrating its neurological effects. Notably, FA is a putative mitochondrial toxin, however, the metabolic effects of FA in the brain remains unknown. Therefore, this study investigates the neurometabolic effects of FA via alterations to Akt and AMPK signalling pathways in C57BL/6 mice at acute (1 day) and prolonged exposure (10 days). Following 1 day exposure, FA augmented Akt signalling by increasing Akt S473 phosphorylation and the upstream regulators PI3K, mTOR and p70S6K. Activated Akt showed inhibition of GSK3 activity with the simultaneous activation of AMPK, p53 phosphorylation and reduced GLUT-1 and -4 receptor expressions, potentially suppressing neuronal glucose entry. However, after 10 days exposure, FA dampened PI3K/Akt and AMPK signalling, but increased the expression of GLUT receptors (1 and 4) in mice brain. Further, FA significantly depleted ATP levels, at 10 days exposure, despite increased PDHE1ß activity (at both 1 and 10 days), strongly suggesting that FA mediates ATP depletion independent of metabolic signalling. In conclusion, FA mediates neurometabolic disturbances, at 1 and 10 day exposures, which may negatively influence normal brain aging and predispose to neurodegenerative disorders.

Mass spectrometric investigations into the brain delivery of abacavir, stavudine and didanosine in a rodent model.

HIV replication in the brain is unopposed due to reduced antiretroviral drug penetration into the central nervous system (CNS). Prevalence of HIV-associated neurocognitive disorder (HAND) has increased severely in patients living with HIV despite current treatments. The aims of this study were to evaluate the brain bio-distribution of alternative nucleoside reverse transcriptase inhibitors, abacavir, stavudine and didanosine in the CNS and to determine their localization patterns in the brain.Sprague-Dawley rats received 50 mg kg-1 single i.p dose of each drug. Mass spectrometric techniques were then used to investigate the pharmacokinetics and localization patterns of these drugs in the brain using LC-MS/MS and mass spectrometric imaging (MSI), respectively.Abacavir, stavudine and didanosine reached the Brain Cmax with concentration of 831.2, 1300 and 43.37 ngmL-1, respectively. Based on MSI analysis Abacavir and Stavudine were located in brain regions that are strongly implicated in the progression of HAND.Abacavir and Stavudine penetrated into CNS, reaching a Cmax that was above the IC50 for HIV (457.6 and 112.0 ngmL-1, respectively), however, it was noted ddI showed poor entry within the brain, therefore, it is recommended that this drug cannot be considered for treating CNS-HIV.

Mass Spectrometry Imaging Demonstrates the Regional Brain Distribution Patterns of Three First-Line Antiretroviral Drugs.

HIV in the central nervous system (CNS) contributes to the development of HIV-associated neurological disorders (HAND), even with chronic antiretroviral therapy. In order for antiretroviral therapy to be effective in protecting the CNS, these drugs should have the ability to localize in brain areas known to be affected by HIV. Consequently, this study aimed to investigate the localization patterns of three first-line antiretroviral drugs, namely, efavirenz, tenofovir, and emtricitabine, in the rat brain. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) were utilized to assess the pharmacokinetics and brain spatial distribution of the three drugs. Each drug was administered (50 mg/kg) to healthy female Sprague-Dawley rats via intraperitoneal administration. LC-MS/MS results showed that all three drugs could be delivered into the brain, although they varied in blood-brain barrier permeability. MALDI-MSI showed a high degree of efavirenz localization across the entire brain, while tenofovir localized mainly in the cortex. Emtricitabine distributed heterogeneously mainly in the thalamus, corpus callosum, and hypothalamus. This study showed that efavirenz, tenofovir, and emtricitabine might be a potential drug combination antiretroviral therapy for CNS protection against HAND.

Investigating time dependent brain distribution of nevirapine via mass spectrometric imaging.

Central nervous system (CNS) HIV infection causes brain tissue inflammation and tissue deficit that contributes to neuroAIDS. This complication is escalated by the blood-brain barrier (BBB), which prevents easy access to antiretroviral drugs entering the CNS. In this study the aims were to investigate the BBB membrane penetration and brain localization patterns of Nevirapine (NVP) using Imaging Mass Spectrometry (MSI). Sprague-Dawley rats received an intraperitoneal dose of NVP (50 mg kg-1). Plasma and brain samples were harvested, and mass spectrometric methods were then applied to determine the pharmacokinetic properties and localization patterns of NVP in the brain. The pharmacokinetic parameters demonstrated a rapid bio-distribution of NVP in plasma and brain. The plasma Cmax was 6320 ng mL-1 and the brain Cmax was 1923 ng mL-1, both at a Tmax of 0.25 h. MSI of coronal brain sections showed that NVP penetrated and localized in the brain regions implicated with the development of HIV associated neurodegeneration. NVP has great potential to combat neuroAIDS.

Zidovudine and Lamivudine as Potential Agents to Combat HIV-Associated Neurocognitive Disorder.

The central nervous system has been identified as an anatomical reservoir for HIV due the difficulties in delivering therapeutic agents into the brain and this complication results in HIV-associated neurocognitive disorder that persists in infected patients. The brain regions that are potentially exposed to tissue deficits due to HIV have been reported in previous reports; therefore, it is important to determine the drugs that can enter and localize in brain regions that are known to be susceptible to HIV neurodegeneration. Sprague-Dawley rats received intraperitoneal doses of zidovudine and lamivudine (50 mg kg-1). Mass spectrometry methods were used to determine the pharmacokinetics, of zidovudine and lamivudine, in the brain using liquid chromatography tandem mass spectrometry and mass spectrometry imaging (MSI), respectively. Zidovudine and lamivudine displayed complementary pharmacokinetic curves indicating a rapid absorption and blood-brain barrier penetration of both drugs reaching Cmaxat 0.5 h after single dose. MSI of coronal brain sections showed that zidovudine and lamivudine are mostly distributed in corpus callosum, globus pallidus, striatum, and the neocortex region. Mass spectrometry techniques were used to demonstrate that zidovudine and lamivudine drugs are able to reach and localize in brain regions that are targets of HIV neurodegeneration in the brain.

Impact of Clofazimine Dosing on Treatment Shortening of the First-Line Regimen in a Mouse Model of Tuberculosis.

The antileprosy drug clofazimine was recently repurposed as part of a newly endorsed short-course regimen for multidrug-resistant tuberculosis. It also enables significant treatment shortening when added to the first-line regimen for drug-susceptible tuberculosis in a mouse model. However, clofazimine causes dose- and duration-dependent skin discoloration in patients, and the optimal clofazimine dosing strategy in the context of the first-line regimen is unknown. We utilized a well-established mouse model to systematically address the impacts of duration, dose, and companion drugs on the treatment-shortening activity of clofazimine in the first-line regimen. In all studies, the primary outcome was relapse-free cure (culture-negative lungs) 6 months after stopping treatment, and the secondary outcome was bactericidal activity, i.e., the decline in the lung bacterial burden during treatment. Our findings indicate that clofazimine activity is most potent when coadministered with first-line drugs continuously throughout treatment and that equivalent treatment-shortening results are obtained with half the dose commonly used in mice. However, our studies also suggest that clofazimine at low exposures may have negative impacts on treatment outcomes, an effect that was evident only after the first 3 months of treatment. These data provide a sound evidence base to inform clofazimine dosing strategies to optimize the antituberculosis effect while minimizing skin discoloration. The results also underscore the importance of conducting long-term studies to allow the full evaluation of drugs administered in combination over long durations.

Toxicity assessment of mycotoxins extracted from contaminated commercial dog pelleted feed on canine blood mononuclear cells.

Raw ingredients of pet food are often contaminated with mycotoxins. This is a serious health problem to pets and causes emotional and economical stress to the pet owners. The aim of this study was to determine the immunotoxicity of the most common mycotoxins (aflatoxin, fumonisin, ochratoxin A and zearalenone) by examining 20 samples of extruded dry dog food found on the South African market [10 samples from standard grocery store lines (SB), 10 from premium veterinarian lines (PB)]. Pelleted dog food was subjected to extraction protocols optimized for the above mentioned mycotoxins. Dog lymphocytes were treated with the extracts (24 h incubation and final concentration 40 µg/ml) to determine cell viability, mitochondrial function, oxidative stress, and markers of cell death using spectrophotometry, luminometry and flow cytometry. Malondialdehyde, a marker of oxidative stress showed no significant difference between SB and PB, however, GSH was significantly depleted in SB extract treatments. Markers of apoptosis (phosphatidylserine externalization) and necrosis (propidium iodide incorporation) were elevated in both food lines when compared to untreated control cells, interestingly SB extracts were significantly higher than PB. We also observed decreased ATP levels and increased mitochondrial depolarization in cells treated with both lines of feed with SB showing the greatest differences when compared to the control. This study provides evidence that irrespective of price, quality or marketing channels, pet foods present a high risk of mycotoxin contamination. Though in this study PB fared better than SB in regards to cell toxicity, there is a multitude of other factors that need to be studied which may have an influence on other negative outcomes.

Clofazimine protects against Mycobacterium tuberculosis dissemination in the central nervous system following aerosol challenge in a murine model.

Tuberculosis (TB) has been the scourge of the human race for many decades, claiming countless number of lives. This is further complicated by the ability of Mycobacterium tuberculosis to infect extrapulmonary sites, specifically the brain. These extrapulmonary forms of TB are difficult to treat owing to problems associated with drug delivery across the blood-brain barrier. Linezolid (LIN) and clofazimine (CFZ) are two of the more promising anti-TB drugs in recent times. In this study, BALB/c mice were aerosol-infected with M. tuberculosis H37Rv and were treated for 4 weeks with LIN [100 mg/kg body weight (BW)] or CFZ (100 mg/kg BW). Concurrently, it was investigated whether an aerosol TB infection would lead to dissemination of TB bacilli into the brain. Post-treatment brain and lung CFUs were determined together with serum, lung and brain drug concentrations. CFZ displayed a strong bactericidal effect in the lung, whilst LIN had a bacteriostatic effect. Mycobacterium tuberculosis appeared at 2 weeks post-infection in the untreated group (2.38 ± 0.43 log10 CFU) and more surprisingly at 3 weeks post-infection in the LIN-treated group (1.14 ± 0.99 log10 CFU). TB bacilli could not be detected in the brains of the CFZ-treated group. To the best of our knowledge, this is the first study showing the appearance of M. tuberculosis in the brain following a murine aerosol TB infection. This study may advocate the use of CFZ as prophylactic treatment to prevent the development of extrapulmonary TB of the central nervous system using a two-pronged approach.

A comparison of mycotoxin contamination of premium and grocery brands of pelleted cat food in South Africa.

Contamination with mycotoxins is of concern to pet owners and veterinary practitioners owing to their ability to cause disease and exacerbate the pathological changes associated with other diseases. Currently, there is a lack of information regarding the mycotoxin content of common premium brand (PB) and grocery brand (GB) cat feeds. Therefore, we undertook to determine the mycobiota content of feed samples, from both categories (n = 6 each), and measured the levels of aflatoxin (AF), fumonisin (FB), ochratoxin A (OTA) and zearalenone (ZEA) by high performance liquid chromatographic analysis. There were high concentrations of mycotoxins in both categories of feed, regardless of the notion that PBs are of a higher quality. The concentration of these toxins may contribute to the development of related pathologies in felines.

A comparative analysis of mycotoxin contamination of supermarket and premium brand pelleted dog food in Durban, South Africa.

Dry pelleted dog food in the South African market is available via supermarkets, pet stores (standard brands [SBs]) and veterinary channels (premium brands [PBs]). For the purpose of this study, the supermarket channel included the cheaper quality foods and PBs were sold via the veterinary channel (n = 20). These feeds were analysed for four main mycotoxins (aflatoxins [AF], fumonisin [FB], ochratoxin A [OTA] and zearalenone [ZEA]) using standard welldescribed extraction, characterisation and quantitation processes. Irrespective of the brand or marketing channel, all foods were contaminated with fungi (mainly Aspergillus flavus, Aspergillus fumigatus and Aspergillus parasiticus) and mycotoxins (most prevalent being aflatoxins and fumonisins). This was observed in all 20 samples irrespective of the marketing channel or perceived quality. Also, many samples within each marketing channel failed the 10 ppb limit for aflatoxin set by regulations in South Africa. Although fumonisin was detected in all samples, a single sample failed the Food and Drug Administration (FDA) limit of 100 ppb. Both OTA and ZEA were found at low concentrations and were absent in some samples. This study suggested that higher priced dog food does not ensure superior quality or that it is free from contamination with fungi or mycotoxins. However, analysis of the more expensive PBs did reveal contamination concentrations lower than those of the SBs.

Lansoprazole-sulfide, pharmacokinetics of this promising anti-tuberculous agent.

Lansoprazole (LPZ) is a commercially available proton-pump inhibitor whose primary metabolite, lansoprazole sulfide (LPZS) was recently reported to have in vitro and in vivo activity against Mycobacterium tuberculosis. It was also reported that a 300 mg kg-1 oral administration of LPZS was necessary to reach therapeutic levels in the lung, with the equivalent human dose being unrealistic. A validated liquid chromatography-tandem mass spectrometric method (LC-MS/MS) for the simultaneous quantification LPZ and LPZS in rat plasma and lung homogenates was developed. We administered 15 mg kg-1 oral doses of LPZ to a healthy rat model to determine the pharmacokinetics of its active metabolite, LPZS, in plasma and lung tissue. We found that the LPZS was present in amounts that were below the limit of quantification. This prompted us to administer the same dose of LPZS to the experimental animals intraperitoneally (i.p.). Using this approach, we found high concentrations of LPZS in plasma and lung, 7841.1 and 9761.2 ng mL-1 , respectively, which were significantly greater than the minimum inhibitory concentration (MIC) for Mycobacterium tuberculosis. While oral and i.p. administration of LPZ resulted in significant concentrations in the lung, it did not undergo sufficient cellular conversion to its anti-TB metabolite. However, when LPZS itself was administered i.p., significant amounts penetrated the tissue. These results have implications for future in vivo studies exploring the potential of LPZS as an anti-TB compound.

Clofazimine has delayed antimicrobial activity against Mycobacterium tuberculosis both in vitro and in vivo.

OBJECTIVES: The anti-leprosy drug clofazimine has been shown to have antimicrobial activity against Mycobacterium tuberculosis and has been associated with treatment-shortening activity in both clinical and preclinical studies of TB chemotherapy. However, a reported lack of early bactericidal activity (EBA) in TB patients has raised questions regarding the usefulness of clofazimine as an anti-TB drug. Our objective was to systematically evaluate the EBA of clofazimine in vitro and in vivo to provide insight into how and when this drug exerts its antimicrobial activity against M. tuberculosis. METHODS: We evaluated the 14 day EBA of clofazimine (i) in vitro at concentrations ranging from 4 times below to 4 times above the MIC for M. tuberculosis and (ii) in vivo in infected BALB/c mice at doses ranging from 1.5 to 100 mg/kg/day, and serum clofazimine levels were measured. In both experiments, isoniazid was used as the positive control. RESULTS: In vitro, clofazimine, at any concentration tested, did not exhibit bactericidal activity during the first week of exposure; however, in the second week, it exhibited concentration-dependent antimicrobial activity. In vivo, clofazimine, at any dose administered, did not exhibit bactericidal activity during the first week, and limited antimicrobial activity was observed during the second week of administration. While serum clofazimine levels were clearly dose dependent, the antimicrobial activity was not significantly related to the dose administered. CONCLUSIONS: Our data suggest that clofazimine's delayed antimicrobial activity may be due more to its mechanism of action rather than to host-related factors.