Toward revealing the role of the cation in the phytotoxicity of the betaine-based esterquats comprising dicamba herbicide.

In this study, two homologous series of esterquats comprising alkyl (from ethyl to octadecyl) betainate cations and bromide as well as dicamba anions were successfully synthesized, starting from a renewable raw material - glycine betaine. Due to the favorable octanol-water partition coefficient and utilization of biodegradable cations of natural origin, synthesized esterquats can be considered promising alternatives to currently applied dicamba-based formulations. In addition, the obtained results allowed us to verify whether the organic cations in quaternary ammonium salts containing herbicidally active anions (such as dicamba) play the role of biologically inactive adjuvants that only enhance the efficiency of the active ingredient or if they simultaneously exhibit a significant degree of phytotoxicity. Analysis of the influence of alkyl betainate esterquats containing nonherbicidal (bromide) anions on seedlings of white mustard revealed that alkyl betainate cations promote the germination of white mustard seeds; however, the subsequent growth of the seedlings was significantly inhibited. Further studies performed on white mustard and cornflower plants in a stage of 4-6 leaves allowed us to conclude that in the case of sensitive plants, the high phytotoxicity can be attributed to the presence of the dicamba anion, whereas for more resistant plants the additional influence of the cation on the phytotoxic effect is visible. Esterquats comprising a dodecyl substituent or longer had high surface active properties. Nonetheless, their contact angle values were not correlated with phytotoxicity data, indicating an additional influence of the cation on this stage of plant development. Interestingly, subsequent dose-response experiments conducted for two selected dicamba-based products confirmed that the greatest phytotoxicity was expressed by compounds containing a decyl substituent.

Predicting the results of a 24-hr human patch test for surfactants: utility of margin-setting in a reconstructed human epidermis model.

To predict the results of a 24-hr closed human patch test, we previously recommended the use of in vitro test with a reconstructed human epidermis (RhE) model adopted in OECD TG 439, and proposed the margin method, which includes evaluation of twice the concentration to avoid a false positive for surfactants. Therefore, in this study, we used LabCyte EPI-MODEL as a RhE model, and confirmed the reproducibility of this method using five surfactants, including benzalkonium chloride (BC), sodium lauryl sulfate (SLS), and lauryl betaine (LB), for which false negative results have previously been reported, and three different surfactants. For all surfactants, prediction of patch test results using a margin of two revealed that human tests could be performed safely, confirming the utility of the margin method. In addition, we examined the relationship with critical micellar concentration (CMC). The IC50 for cell viability in the RhE model for three types of surfactants (BC, SLS, and LB) was 2.7- to 49.7-times the CMC. Therefore, the range of concentrations in which tests were performed with the present method was within the range of concentrations with high cleansing. Furthermore, we examined the relationship between cell viability and release of the inflammatory mediator interleukin-1α (IL-1α). IL-1α release was associated with cell viability, supporting the results of the human patch test.

Biodegradability and aquatic toxicity of new cleavable betainate cationic oligomeric surfactants.

New cleavable oligomeric cationic surfactants containing ester groups susceptible to hydrolysis between the hydrocarbon tails and the hydrophilic moiety have been synthesized and their biodegradability and aquatic toxicity examined. Aerobic biodegradability was evaluated by applying a standard method for ready biodegradability, the CO2 Headspace test. Aquatic toxicity was assessed by means of the acute toxicity test with Daphnia. Cleavable oligomeric cationic surfactants undergo a significant biodegradation extent (31-52%) as compared to dimeric surfactants without ester groups that showed null degradation in previous works. However, they do not attain the threshold of ultimate degradation required (60%) to be classed as easily biodegradable chemicals. On the other hand, the introduction of cleavable groups in the surfactant hydrophobic chains reduces the toxic effects on the microorganisms responsible for degradation observed for conventional alkyl ammonium dimeric surfactants. Acute toxicity values of betainate cationic oligomeric surfactants to Daphnia magna, IC50-48 h, varies from 1.5 to 50 mg/L. Aquatic toxicity of oligomeric cationic surfactants depends on their hydrophobicity and increases regularly with the alkyl chain length. However, whether the surfactant is a dimeric or a trimeric betaine ester does not affect their acute toxicity to crustacean.

Chemometric analysis of Amaranthus retroflexus in relation to livestock toxicity in southern Australia.

Amaranthus retroflexus L., an introduced invasive weed in southern Australia, has been associated with acute renal failure and/or mortality in a number of livestock species. While its leaves, flowers and stems are generally reported to contain high levels of nitrogen, few studies have fully characterised the chemical composition of A. retroflexus foliage with respect to mammalian toxicity. We performed extensive metabolic profiling of stems, leaves, roots and inflorescence tissues of A. retroflexus collected from three spatially and/or temporally distinct toxicity outbreaks, and report on the 1) composition of primary and secondary metabolites in methanolic extracts of A. retroflexus tissues using HPLC and HPLC-MS QToF and 2) chemometric analysis of A. retroflexus extracts in relation to the associated toxin(s). All tissues of A. retroflexus possessed an abundance of N-containing metabolites, particularly quaternary ammonium compounds which were identified as betaines, two of which (valine betaine and isoleucine betaine) are rarely encountered in plants. Cytotoxicity to murine fibroblasts was highest in extracts of leaf tissue and was associated with a single, a small modified peptide with high similarity to N-acetyl-L-α-aspartyl-L-alanyl-L-α-aspartyl-L-α-glutamyl-O-(carboxymethyl)-L-tyrosyl-L-leucinamide, a synthetic phosphotyrosyl mimic involved in cell signaling processes. One possible mode of action leading to acute renal failure in grazing livestock by a modified peptide such as this is proposed.

Mussel-Inspired Surface Functionalization of PET with Zwitterions and Silver Nanoparticles for the Dual-Enhanced Antifouling and Antibacterial Properties.

Herein, we designed and constructed a dual functional surface with antimicrobial and antifouling abilities to prevent protein and bacterial attachment that are significant challenges in biomedical devices. Primary amino-group-capped sulfobetaine of DMMSA was synthesized and then grafted onto polydopamine pretreated PET sheets via click chemistry. The sheets were subsequently immersed into silver ion solution, in which the absorbed silver ions were reduced to silver nanoparticles (AgNPs) in situ by a polydopamine layer. The antifouling assays demonstrated that the resultant PET/DMMSA/AgNPs sheets exhibited great antifouling performances against bovine serum albumin (BSA), bovine fibrinogen (BFG), platelets, and bacteria, the critical proteins/microorganisms leading to implant failure. The antibacterial data suggested that the sheets had dual functions as inhibitors of bacterial growth and bactericide and could efficiently delay the biofilm formation. This repelling and killing approach is green and simple, with potential biomedical applications.

Toxicity of Natural Deep Eutectic Solvent Betaine:Glycerol in Rats.

Natural deep eutectic solvents (NaDES) are new natural solvents in green chemistry that in some cases have been shown to allow better extraction of plant bioactive molecules compared to conventional solvents and higher phenolic compound absorption in rodents. However, there is a serious lack of information regarding their in vivo safety. The purpose of this study was to verify the safety of a NaDES (betaine:glycerol (1:2 mole ratio) of water) extract from green coffee beans, rich in polyphenols. Twelve 6-week-old male Wistar rats were randomized into two groups of 6 animals each and twice daily gavaged for 14 days either with 3 mL of water or 3 mL of phenolic NaDES extract. Oral administration of phenolic NaDES extract induced mortality in two rats. In addition, it induced excessive water consumption, reduced dietary intake and weight loss, hepatomegaly, and plasma oxidative stress associated with high blood lipid levels. In conclusion, this work demonstrated the toxicity of oral administration of the selected NaDES under a short-term condition. This occurs despite the fact that this NaDES extract contains polyphenols, whose beneficial effects have been shown. Therefore, complementary work is needed to find the best dose and formulation of NaDES that are safe for the environment and animals and ultimately for humans.

Enhanced water-solubility, antibacterial activity and biocompatibility upon introducing sulfobetaine and quaternary ammonium to chitosan.

Chitosan (CS) has attracted much attention due to its good antibacterial activity and biocompatibility. However, CS is insoluble in neutral and alkaline aqueous solution, limiting its biomedical application to some extent. To circumvent this drawback, we have synthesized a novel N-quaternary ammonium-O-sulfobetaine-chitosan (Q3BCS) by introducing quaternary ammonium compound (QAC) and sulfobetaine, and its water-solubility, antibacterial activity and biocompatibility were evaluated compare to N-quaternary ammonium chitosan and native CS. The results showed that by introducing QAC, antibacterial activities and water-solubilities increase with degrees of substitution. The largest diameter zone of inhibition (DIZ) was improved from 0 (CS) to 15mm (N-Q3CS). And the water solution became completely transparent from pH 6.5 to pH 11; the maximal waters-solubility was improved from almost 0% (CS) to 113% at pH 7 (N-Q3CS). More importantly, by further introducing sulfobetaine, cell survival rate of Q3BCS increased from 30% (N-Q3CS) to 85% at 2000µg/ml, which is even greater than that of native CS. Furthermore, hemolysis of Q3BCS was dropped sharply from 4.07% (N-Q3CS) to 0.06%, while the water-solution and antibacterial activity were further improved significantly. This work proposes an efficient strategy to prepare CS derivatives with enhanced antibacterial activity, biocompatibility and water-solubility. Additionally, these properties can be finely tailored by changing the feed ratio of CS, glycidyl trimethylammonium chloride and NCO-sulfobetaine.

The in vitro impact of toothpaste extracts on cell viability.

Toothpastes contain three main components: detergents, abrasives, and fluoride. Detergents, particularly sodium lauryl sulfate, have been proposed as components that enable toothpastes to produce cytotoxic effects in vitro. However, not all toothpastes contain sodium lauryl sulfate, and almost no studies have found an association between detergents and the in vitro cytotoxicity of toothpastes. The present study examined the in vitro cytotoxicity of nine commercially available toothpastes containing four different detergents. Toothpastes were diluted in serum-free medium, centrifuged, and filter sterilized. The half-lethal concentration of the toothpaste-conditioned medium (TCM) was calculated based on the formation of formazan by gingival fibroblasts, oral squamous cell carcinoma HSC-2 cells, and L929 cells. Cell proliferation was analyzed, and live-dead staining was performed, after exposure of cells to conditioned medium prepared with 1% toothpaste (1% TCM). It was found that toothpastes containing sodium lauryl sulfate and amine fluoride strongly inhibited cell viability with the half-lethal concentration being obtained with conditioned medium prepared with approximately 1% toothpaste (1% TCM). Toothpastes containing cocamidopropyl betaine and Steareth-20 showed higher half-lethal concentration values, with the half-lethal concentration being obtained with conditioned medium prepared with 10% (10% TCM) and 70% (70% TCM) toothpaste, respectively. Proliferation and live-dead data were consistent with the cell-viability analyses. These results demonstrate that the type of detergent in toothpastes can be associated with changes in in vitro cell toxicity.

Differential immunotoxicities of poly(ethylene glycol)- vs. poly(carboxybetaine)-coated nanoparticles.

Although the careful selection of shell-forming polymers for the construction of nanoparticles is an obvious parameter to consider for shielding of core materials and their payloads, providing for prolonged circulation in vivo by limiting uptake by the immune organs, and thus, allowing accumulation at the target sites, the immunotoxicities that such shielding layers elicit is often overlooked. For instance, we have previously performed rigorous in vitro and in vivo comparisons between two sets of nanoparticles coated with either non-ionic poly(ethylene glycol) (PEG) or zwitterionic poly(carboxybetaine) (PCB), but only now report the immunotoxicity and anti-biofouling properties of both polymers, as homopolymers or nanoparticle-decorating shell, in comparison to the uncoated nanoparticles, and Cremophor-EL, a well-known low molecular weight surfactant used for formulation of several drugs. It was found that both PEG and PCB polymers could induce the expression of cytokines in vitro and in vivo, with PCB being more immunotoxic than PEG, which corroborates the in vivo pharmacokinetics and biodistribution profiles of the two sets of nanoparticles. This is the first study to report on the ability of PEG, the most commonly utilized polymer to coat nanomaterials, and PCB, an emerging zwitterionic anti-biofouling polymer, to induce the secretion of cytokines and be of potential immunotoxicity. Furthermore, we report here on the possible use of immunotoxicity assays to partially predict in vivo pharmacokinetics and biodistribution of nanomaterials.

Betaine attenuates Alzheimer-like pathological changes and memory deficits induced by homocysteine.

Hyperhomocysteinemia (Hhcy) may induce memory deficits with ß-amyloid (Aß) accumulation and tau hyperphosphorylation. Simultaneous supplement of folate and vitamin B12 partially restored the plasma homocysteine level and attenuated tau hyperphosphorylation, Aß accumulation and memory impairments induced by Hhcy. However, folate and vitamin B12 treatment have no effects on Hhcy which has the methylenetetrahydrofolate reductase genotype mutation. In this study, we investigated the effects of simultaneous supplement of betaine on Alzheimer-like pathological changes and memory deficits in hyperhomocysteinemic rats after a 2-week induction by vena caudalis injection of homocysteine (Hcy). We found that supplementation of betaine could ameliorate the Hcy-induced memory deficits, enhance long-term potentiation (LTP) and increase dendritic branches numbers and the density of the dendritic spines, with up-regulation of NR1, NR2A, synaptotagmin, synaptophysin, and phosphorylated synapsin I protein levels. Supplementation of betaine also attenuated the Hcy-induced tau hyperphosphorylation at multiple AD-related sites through activation protein phosphatase-2A (PP2A) with decreased inhibitory demethylated PP2A(C) at Leu309 and phosphorylated PP2A(C) at Tyr307. In addition, supplementation of betaine also decreased Aß production with decreased presenilin-1 protein levels. Our data suggest that betaine could be a promising candidate for arresting Hcy-induced AD-like pathological changes and memory deficits.

Biocompatible zwitterionic sulfobetaine copolymer-coated mesoporous silica nanoparticles for temperature-responsive drug release.

A novel nanocontainer, which could regulate the release of payloads, has been successfully fabricated by attaching zwitterionic sulfobetaine copolymer onto the mesoporous silica nanoparticles (MSNs). RAFT polymerization is employed to prepare the hybrid poly(2-(dimethylamino)ethyl methacrylate)-coated MSNs (MSN-PDMAEMA). Subsequently, the tertiary amine groups in PDMAEMA are quaternized with 1,3-propanesultone to get poly(DMAEMA-co-3-dimethyl(methacryloyloxyethyl)ammonium propanesulfonate)-coated MSNs [MSN-Poly(DMAEMA-co-DMAPS)]. The zwitterionic PDMAPS component endows the nanocarrier with biocompatibility, and the PDMAEMA component makes the copolymer shell temperature-responsive. Controlled release of loaded rhodamine B has been achieved in the saline solutions.

Novas perspectivas de fármacos com propriedade tripanocida: estudos biológicos de compostos heterocíclicos / Novel perspectives of drugs with tripanocidal properties: biological study of heterocyclic compounds

Os compostos mesoiônicos são moléculas não aromáticas planas, pertencentes à classe das betaínas, os quais apresentam um grande número de atividades biológicas incluindo antibacteriana, anti-inflamatória e anti-protozoária. A despeito do potencial quimoterápico destes compostos, estudos destas moléculas contra o Trypanosoma cruzi, agente etiológico da doença de Chagas, ainda são incipientes. No presente trabalho nós avaliamos as atividades tripanocida e citotóxica dos mesoiônicos MECPYR-04, MEC-02, MC-01 e MC-02. Nossos resultados demonstraram que todos os compostos inibiram o crescimento de formas epimastigotas e causaram decréscimo da viabilidade celular em tripomastigotas. Estes compostos também foram efetivos em inibir a infecção de macrófagos e diminuir a viabilidade das formas amastigotas. MECPYR-04 foi o mais efetivo contra todas as formas evolutivas do parasito, entretanto também apresentou alta toxicidade para células de mamíferos, enquanto MEC02, MC-01 e MC-02 apresentaram baixa toxidade em células de mamíferos e alta seletividade para os parasitas. Apenas MEC-02 e MECPYR-04 aumentaram a produção de NO em macrófagos quando comparados com o controle, sugerindo um potencial imunomodulatório destes compostos. Com objetivo de analisar os efeitos de MEC-02, MC-01 e MC-02 sobre a integridade da membrana plasmática e o potencial de membrana mitocondrial, células tratadas e controles foram submetidas a marcação com iodeto de propidio e rodamina 123 respectivamente, e analisadas por microscopia confocal. Nenhuma diferença significativa na integridade da membrana foi observada em células tratadas. Estes achados foram confirmados pela microscopia eletrônica de varredura. Apenas MC-01 e MC-02 foram capazes de induzir alterações no potencial de membrana mitocondrial como observado por microscopia confocal e citometria de fluxo. Análise ultraestrutural revelou apenas pequenas alterações em formas tripomastigotas tratadas com MEC-02. Por outro lado, células tratadas com MC-01 e MC-02 apresentaram drástico inchaço da mitocôndria com desorganização ou perda do material da matriz mitocondrial, sugerindo que esta estrutura seja o alvo de ação de MC-01 e MC-02. Todos estes resultados apontam MEC-02, MC-01 e MC-02 como candidatos a fármacos contra a doença de Chagas.

Toxicity of the amphoteric surfactant, cocamidopropyl betaine, to the marine macroalga, Ulva lactuca.

The degradation of the synthetic, amphoteric surfactant, cocamidopropyl betaine (CAPB) and its toxicity to the marine macroalga, Ulva lactuca, has been evaluated using several different physiological test end-points over different periods of exposure up to 120 h. Droplet surface angle measurements revealed that, following a period of acclimation of about 24 h, CAPB began to degrade and that primary degradation was complete within 120 h. Effective quantum yield (∆F/F(m)') and relative growth rates (RGRs) were the most sensitive measures of phytotoxicity, with CAPB concentrations at and above 10 mg l(-1) eliciting irreversible, time-dependent and/or dose-dependent responses. Cell membrane damage, estimated from measurements of ion leakage, was detected only at a concentration of 40 mg l(-1) after 48 h of exposure to CAPB but by 120 h damage was evident at all measured concentrations above 10 mg l(-1). These observations suggest that both CAPB and its metabolites are intrinsically toxic to U. lactuca. The findings of this study are discussed in terms of the environmental consequences of applying CAPB to control harmful algal blooms.

Modulation of barnacle (Balanus amphitrite Darwin) cyprid settlement behavior by sulfobetaine and carboxybetaine methacrylate polymer coatings.

Zwitterionic polymers such as poly(sulfobetaine methacrylate) (polySBMA) and poly(carboxybetaine methacrylate) (polyCBMA) have demonstrated impressive fouling-resistance against proteins and mammalian cells. In this paper, the effects of these surface chemistries on the settlement and behavior of an ubiquitous fouling organism, the cypris larva of the barnacle Balanus amphitrite (=Amphibalanus amphitrite), were studied in the laboratory. Conventional settlement assays and behavioral analysis of cyprids using Noldus Ethovision 3.1 demonstrated significant differences in settlement and behavior on different surfaces. Cyprids did not settle on the polySBMA or polyCBMA surfaces over the course of the assay, whereas settlement on glass occurred within expected limits. Individual components of cyprid behavior were shown to differ significantly between glass, polySBMA and polyCBMA. Cyprids also responded differently to the two zwitterionic surfaces. On polySBMA, cyprids were unwilling or unable to settle, whereas on polyCBMA cyprids did not attempt exploration and left the surface quickly. In neither case was toxicity observed. It is concluded that a zwitterionic approach to fouling-resistant surface development has considerable potential in marine applications.

Chitosan N-betainates/DNA self-assembly nanoparticles for gene delivery: in vitro uptake and transfection efficiency.

The aim of this work is to investigate the effect of betaine substitution degree of chitosan N-betainates (CsB) on cellular uptake, cytotoxicity and transfection efficiency of CsB/DNA complex nanoparticles (CsBNs) against COS-7 and MDA-MB-468 cells. The polymers with three substitution degrees (CsB12, CsB47 and CsB85) complexed with pDNA formed CsBN12s, CsBN47s and CsBN85s. The CsBNs showed less pH dependency with smaller particle size and higher zeta potential than that of chitosan/pDNA complex nanoparticles (CsNs) at neutral pH. CsBN85s showed stronger cellular uptake than that of CsBN47s or CsBN12s. CsBNs showed higher cytotoxicity than CsNs, and a trend increasing toxicity with substitution degree increasing. In COS-7 cells, the transfection efficiency increased with the substitution degree increasing, while the opposite result was observed in MDA-MB-468 cells. Chitosan modified with betaine could increase its ability to facilitate DNA uptake and its cytotoxicity, both of which showed the influence on transfection efficiency. It was able to increase cellular uptake and transfection efficiency of complex nanoparticles in COS-7 cells to increase betaine substitution of CsB, however, the higher sensitivity of MDA-MB-468 cells to CsBs led to decreased transfection efficiency due to the increased cytotoxicity with betaine substitution increasing. The predominant role of cellular uptake or toxicity in affecting transfection efficiency was different in two cell lines. These results provided an important guidepost for further development of chitosan derivatives/pDNA complexes as non-viral gene vectors.

Ex vivo reconditioning of marginal donor lungs injured by acid aspiration.

BACKGROUND: Injured lungs due to gastric acid aspiration may be rejected for transplantation because of the possibility of early graft dysfunction. We hypothesized that diluted surfactant administration during ex vivo perfusion would recondition the lungs injured by acid aspiration and permit their use as suitable grafts for transplantation. METHODS: Using a pig model, lung injury was induced with 5-ml/kg administration of a betaine-HCl/pepsin mixture via a flexible bronchoscope. After injury, animals were randomly assigned to three study groups (n = 6/group): saline lavage during ex vivo perfusion (control); surfactant lavage ex vivo (SL-Exvivo); and surfactant lavage before harvest (SL-Pre); and a normal group (n = 4), with no lung injury. Cold storage time was 3 hours. A volume of 10 ml/kg (4 mg/ml, 40 mg/kg) surfactant (Curosurf) was used for lavage. Bronchoalveolar lavage (BAL) was performed before and after injury and at the end of the experiment. Protein and neutrophil percentage in BAL were assessed. Hemodynamic and aerodynamic parameters were measured every 30 minutes during a 2-hour observation period. RESULTS: An approximately 50% decrease in Pao(2) was observed in all animals after injury. Ex vivo surfactant lavage resulted in lower pulmonary vascular resistance, lower oxygenation index and higher Pao(2)/Fio(2) ratio compared with the control group (p = 0.001, p = 0.0001 and p = 0.0001, respectively, according to analysis of variance for repeated measures). Wet-to-dry weight ratio was lower in the SL-Exvivo group compared with the control group (p = 0.015). BAL neutrophil percent at the end of the experiment differed significantly between control and all other groups (p < 0.05). CONCLUSION: Diluted surfactant lavage during ex vivo perfusion improves graft function of lungs injured by gastric acid aspiration.

Effect of N-betainate and N-piperazine derivatives of chitosan on the paracellular transport of mannitol in Caco-2 cells.

The effects of novel quaternary chitosan derivatives on the paracellular transport of mannitol and cell viability were studied in the Caco-2 cell model. The N-betainate derivative with the degree of substitution of 0.05 was very effective at 1.0% (w/v) concentration. The activity decreased as the degree of substitution increased. The cytotoxicity of N-betainates was rather low. The N-piperazines were at least equally effective as the N-betainates with a similar degree of substitution (>0.15). Most of the N-piperazines did not exert toxic effects on the cell monolayers. Overall, the inverse proportionality between the degree of substitution and activity suggests that an intact chitosan backbone is essential for the bioactivity of chitosan derivatives. The quaternary group does not substitute for the activity of the free amine group. In particular, the N-betainate derivatives of chitosan should contain only the minimum number of substituents required for water solubility.

Toxicity, inflammation, and anti-human immunodeficiency virus type 1 activity following exposure to chemical moieties of C31G.

C31G, which has potent activity against the human immunodeficiency virus type 1 (HIV-1) and an established record of safety in animal studies and human trials, is a microbicidal agent comprised of a buffered equimolar mixture of two amphoteric, surface-active agents: an alkyl amine oxide (C14AO) and an alkyl betaine (C16B). Studies of long-term in vitro exposure to C31G and its constituents have suggested that the components of C31G may contribute differentially to its toxicity and efficacy. In the present studies, in vitro assays of cytotoxicity and anti-HIV-1 activity demonstrated that C16B was slightly less cytotoxic compared to either C31G or C14AO, whereas the anti-HIV-1 activities of C31G and its individual constituents were similar. In the murine model of cervicovaginal microbicide toxicity, in vivo exposure to C14AO resulted in severe cervical inflammation followed by a delayed disruption of the columnar epithelium. In contrast, exposure to C16B caused severe cervical epithelial disruption and a secondary, less intense inflammatory response. These results demonstrate that (i) there are both mechanistic and temporal differences in toxicity associated with the components of C31G not necessarily predicted by in vitro assessments of cytotoxicity and (ii) contributions of each component to the anti-HIV-1 activity of C31G appear to be equal. In addition, these findings indicate that direct and indirect mechanisms of in vivo toxicity can be observed as separate but interrelated events. These results provide further insight into the activity of C31G, as well as mechanisms potentially associated with microbicide toxicity.

Comparative safety evaluation of the candidate vaginal microbicide C31G.

C31G is currently the focus of clinical trials designed to evaluate this agent as a microbicidal and spermicidal agent. In the following studies, the in vivo safety of C31G was assessed with a Swiss Webster mouse model of cervicovaginal toxicity and correlated with results from in vitro cytotoxicity experiments and published clinical observations. A single exposure of unformulated 1% C31G resulted in mild-to-moderate epithelial disruption and inflammation at 2 and 4 h postapplication. The columnar epithelium of the cervix was the primary site of damage, while no perturbation of the vaginal mucosa was observed. In contrast, application of unformulated 1.7% C31G resulted in greater levels of inflammation in the cervical epithelium at 2 h postapplication and severe epithelial disruption that persisted to 8 h postapplication. Application of a nonionic aqueous gel formulation containing 1% C31G resulted in no apparent cervicovaginal toxicity at any time point evaluated. However, formulation of 1.7% C31G did not substantially reduce the toxicity associated with unformulated C31G at that concentration. These observations correlate with findings gathered during a recent clinical trial, in which once-daily applications resulted in no adverse events in women receiving the formulation containing 1% C31G, compared to moderate-to-severe adverse events in 30% of women receiving the 1.7% C31G formulation. The Swiss Webster mouse model was able to effectively discriminate between concentrations and formulations of C31G that produced distinct clinical effects in human trials. The Swiss Webster animal model may be a highly valuable tool for preclinical evaluation of candidate vaginal microbicides.

Quantitative structure-activity relationship modeling of acute toxicity of quaternary alkylammonium sulfobetaines to Daphnia magna.

The logarithm of the octanol/water partition coefficient has been widely used as the sole physicochemical property to define various types of biological activity, such as aquatic toxicity. In the present study, we derive a quantitative structure-activity relationship (QSAR) equation for the prediction of acute aquatic toxicity (48-h median effective concentration) of quaternary alkylammonium sulfobetaines to the aquatic invertebrate Daphnia magna Straus. The single-parameter approach described is also based on the hydrophobicity parameter, log P. The 17 zwitterionic compounds were considered for QSAR development. They have the general formula RN+(CH3)2(CH2)nSO3-, where n = 2 to 4, and were synthesized by reacting the corresponding N,N-dimethylamines with either sodium-2-chloroethanesulfonate (n = 2), 1,3-propanesultone (n = 3), or 1,4-butanesultone (n = 4). The R group was either a C6 to C12 alkyl chain, a phenylalkyl unit bearing a C1 to C4 chain, or a phenylpropyl unit with a C4 to C6 para-substituted alkyl group. Octanol/water partition coefficients were experimentally determined via a conventional stir-flask procedure, which was quantified using a reverse-phase, high-performance liquid chromatographic technique coupled with either an ultraviolet or an electrospray ionization mass spectrometry detection system. Median effective concentration values were also determined experimentally via a standard acute immobilization test recommended by Organisation for Economic Cooperation and Development (Paris, France) Guideline 202. The established QSAR equation for such zwitterionic compounds is consistent with the standard equation that normally defines the polar narcosis mode of toxic action.