Ruminal modulator additive effect of Stryphnodendron rotundifolium bark in feedlot lambs.

The study aimed to evaluate the inclusion effects of Stryphnodendron rotundifolium (barbatimão) extracts in substitution of the lasalocid sodium on the ingestive behaviour, intake, ruminal parameters, and digestibility of feedlot lambs. Twenty-four pantaneiro lambs were used, with an average age of 150 ± 4.59 days and an initial body weight of 21.2 ± 3.63 kg. The lambs were distributed in three treatments in an experimental design with randomized blocks. The treatments correspond to the additive supplements: LAS (0.019 g of lasalocid sodium/lamb/d); DGB (1.50 g of barbatimão dried ground bark/lamb/d); DHE (0.30 g of barbatimão dry hydroalcoholic extract/lamb/d). The DHE increased 59.74 min in the time spent for ingestion per day, resulting in an efficiency reduction of dry matter (DM) ingestion (127 g of DM/h of feed). There was a reduction of 1.8 mg/dL in the ammoniacal nitrogen concentration with extract supplementation compared to LAS. The DGB reduced total volatile fatty acids by 48.9% compared to the control treatment. The inclusion of barbatimão extracts (DGB and DHE) reduced 12.05% of ruminal butyrate content. The supplementation of barbatimão extracts replacing lasalocid sodium in the diet of feedlot lambs did not affect intake and caused small changes on ingestive behaviour.

Simultaneous Improvement in the Thermostability and Catalytic Activity of Epoxidase Lsd18 for the Synthesis of Lasalocid A.

Enzymes used in the synthesis of natural products are potent catalysts, capable of efficient and stereoselective chemical transformations. Lsd18 catalyzes two sequential epoxidations during the biosynthesis of lasalocid A, a polyether polyketide natural product. We performed protein engineering on Lsd18 to improve its thermostability and catalytic activity. Utilizing structure-guided methods of FoldX and Rosetta-ddG, we designed 15 mutants of Lsd18. Screening of these mutants using thermal shift assay identified stabilized variants Lsd18-T189M, Lsd18-S195M, and the double mutant Lsd18-T189M-S195M. Trypsin digestion, molecular dynamic simulation, circular dichroism (CD) spectroscopy, and X-ray crystallography provided insights into the molecular basis for the improved enzyme properties. Notably, enhanced hydrophobic interaction within the enzyme core and interaction of the protein with the FAD cofactor appear to be responsible for its better thermostability.

New Hydrophilic Derivatives of Lasalocid and Their Complexes with Selected Metal Cations.

Two new esters of lasalocid, that are more hydrophilic, with glucose (LasGlu) and xylitol (LasX), have been synthesized, and their complexation of monovalent cations has been studied by various spectrometric and spectroscopic methods, such as ESI mass spectrometry, 1H, 13C NMR and FT-IR. Analyses of the results confirmed the synthesis of new esters with good yields. In order to carry out further studies, it was necessary to purify them using "flash" liquid chromatography. It was confirmed that the newly obtained molecules, as well as their complexes with lithium, sodium and potassium cations, were stabilized by a strong system of intramolecular hydrogen bonds. It was found that the hydroxyl groups of esters derived from xylitol and glucose were also involved in the complexation of cations. The results of the PM6 semiempirical calculations permitted determination of the heat of formation (HOF), and visualization of the structure of the new esters and their complexes with the cations studied. All computation results are in agreement with the spectroscopic data.

Analysis of electrochemical and liver microsomal transformation products of lasalocid by LC/HRMS.

RATIONALE: Lasalocid (LAS), an ionophore, is used in cattle and poultry farming as feed additive for its antibiotic and growth-promoting properties. Literature on transformation products (TP) resulting from LAS degradation is limited. So far, only hydroxylation is found to occur as the metabolic reaction during the LAS degradation. To investigate potential TPs of LAS, we used electrochemistry (EC) and liver microsome (LM) assays to synthesize TPs, which were identified using liquid chromatography high-resolution mass spectrometry (LC/HRMS). METHODS: Electrochemically produced TPs were analyzed online by direct coupling of the electrochemical cell to the electrospray ionization (ESI) source of a Sciex Triple-TOF high resolution mass spectrometer. Then, EC-treated LAS solution was collected and analyzed offline using LC/HRMS to confirm stable TPs and improve their annotation with a chemical structure due to informative MS/MS spectra. In a complementary approach, TPs formed by rat and human microsomal incubation were investigated using LC/HRMS. The resulting data were used to investigate LAS modification reactions and elucidate the chemical structure of obtained TPs. RESULTS: The online measurements identified a broad variety of TPs, resulting from modification reactions like (de-)hydrogenation, hydration, methylation, oxidation as well as adduct formation with methanol. We consistently observed different ion complexations of LAS and LAS-TPs (Na+ ; 2Na+ K+ ; NaNH4 + ; KNH4 + ). Two stable methylated EC-TPs were found, structurally annotated, and assigned to a likely modification reaction. Using LM incubation, seven TPs were formed, mostly by oxidation/hydroxylation. After the identification of LM-TPs as Na+ -complexes, we identified LM-TPs as K+ -complexes. CONCLUSION: We identified and characterized TPs of LAS using EC- and LM-based methods. Moreover, we found different ion complexes of LAS-based TPs. This knowledge, especially the different ion complexes, may help elucidate the metabolic and environmental degradation pathways of LAS.

Detection of salinomycin and lasalocid in chicken liver by icELISA based on functional bispecific single-chain antibody (scDb) and interpretation of molecular recognition mechanism.

Salinomycin (SAL) and lasalocid (LAS) are widely used as ionophore antibiotics for coccidiosis control. However, their common use as feed additives has led to the occurrence of feed cross-contamination, which has toxic effects on non-target animals. There have been few reports on multiple-residue detection for SAL and LAS in recent years. In this study, two single-chain antibody fragments (scFvs) capable of specifically recognizing SAL and LAS were constructed. Using LAS-scFv and SAL-scFv as parent antibodies, a complete bispecific single-chain diabody (scDb) against both LAS and SAL was built using splicing by overlap extension polymerase chain reaction (SOE-PCR). In addition, the key amino acid sites and interaction energy of antibody variable regions for small-molecule recognition were preliminarily studied by homology modeling and molecular docking. Finally, IC50 values of 12.9 and 8.6 ng/mL, with a linear range of 6.9-24.0 and 4.7-16.0 ng/mL, were obtained for LAS-scFv and SAL-scFv, respectively. An indirect competitive enzyme-linked immunosorbent assay (icELISA) method was established using scDb to obtain an IC50 of 3.5 ng/mL for LAS and 4.1 ng/mL for SAL, which showed better sensitivity and specificity than those of the parent scFv antibodies. The recoveries of LAS and SAL in chicken liver were 89.2-92.7%(CV<4.7%) and 88.6-90.2% (CV<6.8%)), respectively.

Lasalocid Acid Antibiotic at a Membrane Surface Probed by Sum Frequency Generation Spectroscopy.

Carboxyl polyether ionophores (CPIs) are widely used as veterinary antibiotics and to increase food utilization in ruminating animals. Furthermore, CPIs can target drug-resistant bacteria, but detailed knowledge about their mode-of-action is needed to develop agents with a reasonable therapeutic index. It has been suggested that ionophores bind to membranes and incur large structural changes to shield a bound ion from the hydrophobic environment of the lipid bilayer for transport. One crucial piece of information is missing, however: Is it necessary for the free ionophore to adsorb on the membrane surface before interacting with a cation to facilitate cross-membrane ion transport? To answer this question, we applied sum-frequency generation (SFG) vibrational spectroscopy and surface tensiometry to identify the interaction between the prototypical CPI lasalocid acid (LA) and a model membrane. Observed changes in the surface pressure demonstrate that the free LA undergoes a self-assembly process with the lipid monolayer. Spectra taken from the lipid monolayer show that the free acid inserts partially into the lipid monolayer and then after complexation with sodium chloride disrupts the lipid monolayer. Overall, this study strongly suggests that this must be the crucial step of LA and metal ion complexation that allows the ionophore to traverse a lipid membrane.

Streptomyces lasalocidi sp. nov. (formerly 'Streptomyces lasaliensis'), an actinomycete isolated from soil which produces the polyether antibiotic lasalocid.

Strain ATCC 31180T was isolated from soil collected in Hyde Park, Massachusetts (USA), and found to produce the polyether antibiotic lasalocid. The name 'Streptomyces lasaliensis' has been in common use since 1974, without a recognized taxonomic description. The most closely related type cultures determined by rRNA gene sequence similarity were Streptomyces longwoodensis DSM 41677T (100 %) and Streptomyces galbus DSM 40089T (100 %). OrthoANI values with S. longwoodensis and S. galbus were 95.50 and 94.41 %, respectively. Chemotaxonomic characteristics supported inclusion within the genus Streptomyces. The cell wall peptidoglycan contained ll-diaminopimelic acid, and the major whole-cell sugars were glucose and ribose. Polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylinositol, phosphatidylglycerol, one unidentified lipid and one unidentified glycolipid. The major menaquinones detected were MK9(H4), MK9(H6) and MK9(H8). The major cellular fatty acids were anteiso-C15 : 0, anteiso-C17 : 0, iso-C16 : 0, iso-C15 : 0 and anteiso-C17 : 1. Its DNA had a G+C content of 72.6 %. Differentiation of ATCC 31180T from the closely related species was evident from digital DNA-DNA hybridization values of 61.80 and 56.90 % for S. longwoodensis and S. galbus respectively. Significant differences were seen in the polyphasic phenotypic analyses. ATCC 31180T produced lasalocid, grew from 10 to 45 °C, pH4-8 and in the presence of 0-10 % NaCl, 0.01 % NaN3 and 1 % phenol. Melanin was produced; H2S and indole were not. Nitrate was not reduced. Spore chains were retinaculum-apertum and spore surfaces were smooth. Spore colour, mycelia colour and soluble pigment production were medium-dependent. The proposed name is Streptomyces lasalocidi sp. nov.; the type strain being ATCC 31180T (=NRRL 3382T=DSM 46487T).

Determination of Eight Coccidiostats in Eggs by Liquid-Liquid Extraction-Solid-Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry.

A method for the simultaneous determination of robenidine, halofuginone, lasalocid, monensin, nigericin, salinomycin, narasin, and maduramicin residues in eggs by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. The sample preparation method used a combination of liquid-liquid extraction (LLE) and solid-phase extraction (SPE) technology to extract and purify these target compounds from eggs. The target compounds were separated by gradient elution using high-performance liquid chromatography (HPLC) and ultra-performance liquid chromatography (UPLC). Tandem mass spectrometry was used to quantitatively and qualitatively analyze the target compounds via electrospray ionization (ESI+) and multiple reaction monitoring mode. The HPLC-MS/MS and UPLC-MS/MS methods were validated according to the requirements defined by the European Union and the Food and Drug Administration. The limits of detection and limits of quantification of the eight coccidiostats in eggs were 0.23-0.52 µg/kg and 0.82-1.73 µg/kg for HPLC-MS/MS, and 0.16-0.42 µg/kg and 0.81-1.25 µg/kg for UPLC-MS/MS, respectively. The eggs were spiked with four concentrations of the eight coccidiostats, and the HPLC-MS/MS and UPLC-MS/MS average recoveries were all higher than 71.69% and 72.26%, respectively. Compared with the HPLC-MS/MS method, utilizing UPLC-MS/MS had the advantages of low reagent consumption, a short detection time, and high recovery and precision. Finally, the HPLC-MS/MS and UPLC-MS/MS methods were successfully applied to detect eight coccidiostats in 40 eggs.

Lasalocid immediately and completely prevents the myocardial damage caused by coronary ischemia reperfusion in rat heart.

Lasalocid, a specific mobile membrane ionophore for calcium, dopamine and norepinephrine was assayed in its capacity to reduce or maintain unaltered the cardiovascular function in conditions of imminent myocardial injury. In experiments of coronary blockade and reperfusion carried out in rat heart, it was found that when administered from 5 to 30 minutes prior to the induction of coronary blockade, at a concentration of 2 mg/kg of body weight, the ionophore immediately, simultaneously, and completely interrupts the blood pressure decay, cardiac frequency increase, electrical ventricular tachycardia and fibrillation, as well as the fall of mitochondrial oxidative phosphorylation and decay of mitochondrial oxygen uptake provoked by the induced myocardial injury. It appears that the molecular mode of action of the lasalocid is associated with its unique ability to transport both calcium and the catecholamines, dopamine and norepinephrine, across mitochondrial and bimolecular lipid membranes, as well as through synaptic cell membrane terminals from rat heart, myocardial fibers of the heart and heart chromaffin membrane vesicles. It is suggested that for the potential medical use of lasalocid to detain incoming ischemic myocardial damage, there exists a need to develop a personal electronic device able to simultaneously monitor, detect, and inform on the very early and simultaneous signs of cardiac alterations of electrical, mechano-chemical, metabolic and hydraulic nature, all which precede heart failure and to administer the lasalocid.

Enzymatic catalysis of anti-Baldwin ring closure in polyether biosynthesis.

Polycyclic polyether natural products have fascinated chemists and biologists alike owing to their useful biological activity, highly complex structure and intriguing biosynthetic mechanisms. Following the original proposal for the polyepoxide origin of lasalocid and isolasalocid and the experimental determination of the origins of the oxygen and carbon atoms of both lasalocid and monensin, a unified stereochemical model for the biosynthesis of polyether ionophore antibiotics was proposed. The model was based on a cascade of nucleophilic ring closures of postulated polyepoxide substrates generated by stereospecific oxidation of all-trans polyene polyketide intermediates. Shortly thereafter, a related model was proposed for the biogenesis of marine ladder toxins, involving a series of nominally disfavoured anti-Baldwin, endo-tet epoxide-ring-opening reactions. Recently, we identified Lsd19 from the Streptomyces lasaliensis gene cluster as the epoxide hydrolase responsible for the epoxide-opening cyclization of bisepoxyprelasalocid A to form lasalocid A. Here we report the X-ray crystal structure of Lsd19 in complex with its substrate and product analogue to provide the first atomic structure-to our knowledge-of a natural enzyme capable of catalysing the disfavoured epoxide-opening cyclic ether formation. On the basis of our structural and computational studies, we propose a general mechanism for the enzymatic catalysis of polyether natural product biosynthesis.

Repurposing Ionophores as novel antimicrobial agents for the treatment of bovine mastitis caused by Gram-positive pathogens.

Increasing reports of multidrug-resistant bacterial infections in animals has created a need for novel antimicrobial agents that do not promote cross-resistance to critically important antimicrobial classes used in human medicine. In response to the recent emergence of antimicrobial resistance in several bovine mastitis pathogens, in vitro antimicrobial susceptibility was determined for four polyether ionophores (lasalocid, monensin, narasin and salinomycin) against Staphylococcus spp. and Streptococcus spp. isolated from clinical cases. In addition, erythrocyte haemolysis and WST-1 cell proliferation assays were used to assess in vitro mammalian cell cytotoxicity and biofilm susceptibility testing was performed using the minimum biofilm eradication concentration (MBEC™) biofilm assay. Lasalocid, monensin, narasin and salinomycin exhibited bacteriostatic antimicrobial activity against all pathogens tested, including methicillin-resistant staphylococci, with MIC90 values <16 µg/ml. Narasin and monensin displayed the least toxicity against mammalian cell lines and all compounds significantly reduced viable cell numbers in a Staphylococcus aureus biofilm. Based on in vitro characterization, all four ionophores offer potentially novel treatments against bovine mastitis but in vivo studies will be essential to determine whether acceptable safety and efficacy is present following intramammary administration.

Trypanosoma brucei: trypanocidal and cell swelling activities of lasalocid acid.

Chemotherapeutic treatment of human and animal trypanosomiasis is unsatisfactory because only a few drugs are available. As these drugs have poor efficacy and cause adverse reactions, more effective and tolerable medications are needed. As the polyether ionophore antibiotic lasalocid acid is used as medicated feed additive in cattle, the compound was tested for its trypanocidal and cytotoxic activity against bloodstream forms of Trypanosoma brucei and human myeloid HL-60 cells. The concentrations required of lasalocid acid to reduce the growth rate of trypanosomes by 50% and to kill the parasites were 1.75 and 10 µM, respectively. The ionophore displayed also cytotoxic activity against HL-60 cells but the human cells were about 10 to 14 times less sensitive indicating moderate selectivity. As the trypanocidal mechanism of action of polyether ionophore antibiotics is due to a sodium influx-induced cell swelling, the effect of lasalocid acid on cell volume change in bloodstream-form trypanosomes was investigated. Interestingly, lasalocid acid induced a much faster cell swelling in trypanosomes than the more trypanocidal related ionophore salinomycin. These results support further investigations of lasalocid acid and derivatives thereof as potential agents against African trypanosomiasis.

In vitro comparison of the dermal penetration of three different topical formulations containing lasalocid.

BACKGROUND: Topical antimicrobial preparations are of utmost importance in treating suspected and confirmed meticillin-resistant Staphylococcus pseudintermedius (MRSP) infections due to the increasing incidence of widespread resistance to systemic antimicrobials. Lasalocid is active against MRSP in vitro and this may become an important topical antimicrobial for the treatment of canine pyoderma. HYPOTHESIS/OBJECTIVES: To determine effects of various formulation types on penetration and retention of lasalocid applied to canine skin in vitro. ANIMALS: Normal canine skin was collected from the thorax of five dogs that had been euthanized on the basis of health and/or intractable behavioural issues. METHODS: Solution, lotion and ointment containing 2% lasalocid were applied to ex vivo canine skin. Transdermal penetration was assessed for a 24 h period and retention of lasalocid was assessed at the conclusion of the study. RESULTS: The solution had significantly higher skin retention of lasalocid and proportion of applied dose retained in skin than lotion and ointment (Tukey-Kramer Honest Significant Difference test, P < 0.01). Lasalocid could not be detected in the receptor fluid of any Franz cell at any time point. CONCLUSIONS AND CLINICAL IMPORTANCE: Lasalocid was not identified in the receptor fluid of any sample, indicating that systemic absorption of the active ingredient in vivo is unlikely. Lasalocid may be useful in the treatment of MRSP infections if in vivo studies support safety and efficacy.

Performance of beef heifers supplemented with sodium lasalocid.

This study was conducted on 78 13-month-old crossbred beef heifers that weighed 215 kg in Southern Rio Grande do Sul (RS) State, Brazil. We evaluated the performance of beef heifers that were reared in a pasture system that received a mineral supplement energy-type protein with added sodium lasalocid (LAS). The heifers were randomly and uniformly divided into 2 groups, with 39 animals in each group. One group of animals received a mineral supplement energy-type protein without sodium lasalocid (CON), and the other group received a mineral supplement energy-type protein with added LAS. The mean feed intake, the body weight (BW), the average daily gain (ADG), the body condition score (BCS), and ovarian cyclicity were recorded, and economic parameters were calculated. No differences in supplement intake were observed between the groups, which ensures adequate intake of the other components of the mineral mixture, which are part of the nutritional requirements for the production process. Similarly, no difference in the ADG was observed between treatments. We observed that the heifers in the LAS group had a higher BW gain (51 kg) that the CON heifers (40 kg; P < 0.05). In addition, LAS-supplemented heifers had a higher BCS (3.53) than CON heifers (3.38) at the end of the experiment (P < 0.05). The heifers supplemented with LAS had a higher profitability than the CON heifers, even with the higher cost of the supplement containing LAS; this effect was due to the higher live BW at the end of the study. We concluded that the administration of a mineral supplement energy-type protein with added LAS has beneficial effects on beef heifers in terms of production and economic feasibility.

Evaluation of certain veterinary drug residues in food. Eighty-first report of the Joint FAO/WHO Expert Committee on Food Additives.

This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of residues of certain veterinary drugs in food and to recommend maximum levels for such residues in food. The first part of the report considers general principles regarding the evaluation of residues of veterinary drugs within the terms of reference of the Joint FAO/WHO Expert Committee on Food Additives (JECFA), including MRLs for generic fish species, acute reference doses (ARfDs) for veterinary drugs, an approach for dietary exposure assessment of compounds used for multiple purposes (i.e veterinary drugs and pesticides), dietary exposure assessment for less-than-lifetime exposure, and the assessment of short-term (90-day and 12-month) studies in dogs. Summaries follow of the Committee's evaluations of toxicological and residue data on a variety of veterinary drugs: two insecticides (diflubenzuron and teflubenzuron), an antiparasitic agent (ivermectin), an ectoparasiticide (sisapronil) and a ß2-adrenoceptor agonist (zilpaterol hydrochloride). In addition, the Committee considered issues raised in concern forms from the Codex Committee on Residues of Veterinary Drugs in Foods on lasalocid sodium, an antiparasitic agent. Annexed to the report is a summary of the Committee's recommendations on these drugs, including acceptable daily intakes (ADIs), ARfDs and proposed MRLs.

Epoxide hydrolase-lasalocid a structure provides mechanistic insight into polyether natural product biosynthesis.

Biosynthesis of some polyether natural products involves a kinetically disfavored epoxide-opening cyclic ether formation, a reaction termed anti-Baldwin cyclization. One such example is the biosynthesis of lasalocid A, an ionophore antibiotic polyether. During lasalocid A biosynthesis, an epoxide hydrolase, Lsd19, converts the bisepoxy polyketide intermediate into the tetrahydrofuranyl-tetrahydropyran product. We report the crystal structure of Lsd19 in complex with lasalocid A. The structure unambiguously shows that the C-terminal domain of Lsd19 catalyzes the intriguing anti-Baldwin cyclization. We propose a general mechanism for epoxide selection by ionophore polyether epoxide hydrolases.

Assessment of probiotics supplementation via feed or water on the growth performance, intestinal morphology and microflora of chickens after experimental infection with Eimeria acervulina, Eimeria maxima and Eimeria tenella.

In this study, the effect of probiotic supplementation via drinking water or feed on the performance of broiler chickens experimentally infected with sporulated oocysts of Eimeria acervulina (5 × 10(4)), Eimeria maxima and Eimeria tenella (2 × 10(4) each one) at 14 days of age was evaluated. Two hundred and forty 1-day-old Ross 308 male chicks were separated into eight equal groups with three replicates. Two of the groups, one infected with mixed Eimeria oocysts and the other not, were given a basal diet and served as controls. The remaining groups were also challenged with mixed Eimeria species and received the basal diet and either water supplemented with probiotic (three groups) or probiotic via feed (two groups); the probiotic used consisted of Enterococcus faecium #589, Bifidobacterium animalis #503 and Lactobacillus salivarius #505 at a ratio of 6:3:1. Probiotic supplementation was applied either via drinking water in different inclusion rates (groups W1, W2 and W3) or via feed using uncoated (group FN) or coated strains (group FC). The last group was given the basal diet supplemented with the anticoccidial lasalocid at 75 mg/kg. Each experimental group was given the corresponding diet or drinking water from day 1 to day 42 of age. Throughout the experimental period of 42 days, body weight and feed intake were recorded weekly and feed conversion ratios were calculated. Seven days after infection, the infected control group presented the lowest weight gain values, while probiotics supplied via feed supported growth to a comparable level with that of the lasalocid group. Probiotic groups presented lesion score values and oocyst numbers that were lower than in control infected birds but higher than in the lasalocid group. In the duodenum, jejunum and ileum, the highest villous height values were presented by probiotic groups. In conclusion, a mixture of probiotic substances gave considerable improvement in both growth performance and intestinal health in comparison with infected control birds and fairly similar improvement to an approved anticoccidial during a mixed Eimeria infection.

Population Pharmacokinetics and Absolute Oral Bioavailability of Lasalocid after Single Intravenous and Intracrop Administration in Laying Hens.

Lasalocid sodium is a polyether carboxylic ionophore agent authorized by the EU for use as a coccidiostat in broilers, turkeys, and pullets up to 16 weeks of age, except for laying hens. However, laying hens are the most common nontarget species exposed to lasalocid sodium, mainly due to cross-contamination from feed mills. This exposure may result in potential drug residue deposition in eggs, which could potentially expose consumers to the drug. The breeds commonly used for commercial egg production in Poland are Isa Brown and Green-legged Partridge hens, which have been found to significantly differ in egg-laying performance. This variability may also affect the pharmacokinetics of lasalocid. Data on lasalocid plasma pharmacokinetics in laying hens are lacking. In this study, we aimed to determine typical population pharmacokinetic parameters, absolute oral bioavailability, and how breed may influence the pharmacokinetics of lasalocid. Twenty-layer hens of the two breeds were used in this study. Lasalocid was administered orally at a single dose of either 1 mg or 5 mg/kg body weight or intravenously at a dose of 1 mg/kg body weight, in a crossover design with a three-week washout period between study periods. Blood samples were collected for 72 h, and lasalocid concentrations were measured using high-performance liquid chromatography with fluorescence detection. A population pharmacokinetic analysis was conducted using nonlinear mixed effects modeling. Standard numerical and graphical criteria were used to select the best model, and a stepwise covariate modeling approach was used to determine any influencing factors. The best model was a three-compartment mammillary model with first-order absorption, transit compartments, and linear elimination. The estimated absolute oral bioavailability was low (36%). It was found that breed significantly influenced not only absorption but also the elimination of lasalocid. This study revealed that lasalocid absorption and elimination varied between the two breeds. This variability in pharmacokinetics may result in breed-related differences in drug residue accumulation in eggs, and ultimately, the risk associated with consumer exposure to drug residues may also vary.

Ultrastructural changes in cardiac and skeletal myoblasts following in vitro exposure to monensin, salinomycin, and lasalocid.

Carboxylic ionophores are polyether antibiotics used in production animals as feed additives, with a wide range of benefits. However, ionophore toxicosis often occurs as a result of food mixing errors or extra-label use and primarily targets the cardiac and skeletal muscles of livestock. The ultrastructural changes induced by 48 hours of exposure to 0.1 µM monensin, salinomycin, and lasalocid in cardiac (H9c2) and skeletal (L6) myoblasts in vitro were investigated using transmission electron microscopy and scanning electron microscopy. Ionophore exposure resulted in condensed mitochondria, dilated Golgi apparatus, and cytoplasmic vacuolization which appeared as indentations on the myoblast surface. Ultrastructurally, it appears that both apoptotic and necrotic myoblasts were present after exposure to the ionophores. Apoptotic myoblasts contained condensed chromatin and apoptotic bodies budding from their surface. Necrotic myoblasts had disrupted plasma membranes and damaged cytoplasmic organelles. Of the three ionophores, monensin induced the most alterations in myoblasts of both cell lines.

One-pot synthesis and cytotoxicity studies of new Mannich base derivatives of polyether antibiotic--lasalocid acid.

Seven Mannich base derivatives of polyether antibiotic Lasalocid acid (2a-2g) were synthesized and screened for their antiproliferative activity against various human cancer cell lines. A novel chemoselective one-pot synthesis of these Mannich bases was developed. Compounds 2a-2c and 2g with sterically smaller dialkylamine substituent, displayed potent antiproliferative activity (IC50: 3.2-7.3 µM), and demonstrated higher than twofold selectivity for specific type of cancer. The nature of Mannich base substituent on C-2 atom at the aromatic ring may be critical in the search for selectivity towards a particular cancer cell.