Ability of biomimetic chromatography and physicochemical systems to predict the skin permeation of neutral compounds. A comparison study.

Five in vitro physicochemical systems have been evaluated in terms of its ability to emulate the skin permeation of neutral compounds: the permeation in two different PAMPA membranes, the classical octanol-water partition coefficient, and two biomimetic chromatography systems, one based in cerasome electrokinetic chromatography and another based in reversed-phase liquid chromatography measurements. The coefficients of the solvation parameter model equation of the mentioned systems have been compared to the ones of the skin permeation process through different comparison parameters. Moreover, a method to predict whether a physicochemical system is able to emulate satisfactorily a biological one, just by the analysis of the equation coefficients has been developed. Results reveal that the two PAMPA systems are a good choice to emulate directly the skin permeation of neutral compounds. Instead, the other three systems need a volume correction term to provide a satisfactory emulation. However, after the correction, all the evaluated systems show a similar ability to emulate well skin permeation, as predicted.

Collection of Partition Coefficients in Hexadecyltrimethylammonium Bromide, Sodium Cholate, and Lithium Perfluorooctanesulfonate Micellar Solutions: Experimental Determination and Computational Predictions.

This study focuses on determining the partition coefficients (logP) of a diverse set of 63 molecules in three distinct micellar systems: hexadecyltrimethylammonium bromide (HTAB), sodium cholate (SC), and lithium perfluorooctanesulfonate (LPFOS). The experimental log p values were obtained through micellar electrokinetic chromatography (MEKC) experiments, conducted under controlled pH conditions. Then, Quantum Mechanics (QM) and machine learning approaches are proposed for the prediction of the partition coefficients in these three micellar systems. In the applied QM approach, the experimentally obtained partition coefficients were correlated with the calculated values for the case of the 15 solvent mixtures. Using Density Function Theory (DFT) with the B3LYP functional, we calculated the solvation free energies of 63 molecules in these 16 solvents. The combined data from the experimental partition coefficients in the three micellar formulations showed that the 1-propanol/water combination demonstrated the best agreement with the experimental partition coefficients for the SC and HTAB micelles. Moreover, we employed the SVM approach and k-means clustering based on the generation of the chemical descriptor space. The analysis revealed distinct partitioning patterns associated with specific characteristic features within each identified class. These results indicate the utility of the combined techniques when we want an efficient and quicker model for predicting partition coefficients in diverse micelles.

Evaluation of the Ability of PAMPA Membranes to Emulate Biological Processes through the Abraham Solvation Parameter Model.

Two parallel artificial membrane permeability assay (PAMPA) systems intended for emulating skin permeability have been characterized through the solvation parameter model of Abraham using multilinear regression analysis. The coefficients of the obtained equations have been compared to the ones already established for other PAMPA membranes using statistical tools. The results indicate that both skin membranes are similar to each other in their physicochemical properties. However, they are different from other PAMPA membranes (e.g., intestinal absorption and blood-brain PAMPAs), mainly in terms of hydrophobicity and hydrogen bonding properties. Next, all PAMPA membranes have been compared to relevant biological processes also characterized through the solvation parameter model. The results highlight that skin-PAMPA membranes are a very good choice to emulate skin permeability.

Virtual Cocrystal Screening of Adefovir Dipivoxyl: Identification of New Solid Forms with Improved Dissolution and Permeation Profiles.

The application of a computational screening methodology based on the calculation of intermolecular interaction energies has guided the discovery of new multicomponent solid forms of the oral antiviral Adefovir Dipivoxyl. Three new cocrystals with resorcinol, orcinol and hydroquinone have been synthesized and thoroughly characterized. They show improved dissolution profiles with respect to the single solid form, particularly the cocrystals of orcinol and resorcinol, which have 3.2- and 2-fold faster dissolution rates at stomach conditions (pH 1.5). Moreover, dynamic dissolution experiments that simultaneously mimic both the pH variation along the gastrointestinal tract and the partition into biological membranes show that, in addition to the faster initial dissolution, Adefovir Dipivoxyl also penetrates faster into the organic membranes in the form of resorcinol and orcinol cocrystals.

Determination of the aqueous pKa of very insoluble drugs by capillary electrophoresis: Internal standards for methanol-water extrapolation.

A fast determination of acidity constants (pKa) of very insoluble drugs has become a necessity in drug discovery process because it often produces molecules that are highly lipophilic and sparingly soluble in water. In this work the high throughput internal standard capillary electrophoresis (IS-CE) method has been adapted to the determination of pKa of water insoluble compounds by measurement in methanol/aqueous buffer mixtures. For this purpose, the reference pKa values for a set of 46 acid-base compounds of varied structure (internal standards) have been established in methanol-water mixtures at several solvent composition levels (with a maximum of 40% methanol). The IS-CE method has been successfully applied to seven test drugs of different chemical nature with intrinsic solubilities lower than 10-6 M. pKa values have been determined at different methanol/aqueous buffer compositions and afterwards Yasuda-Shedlovsky extrapolation method has been applied to obtain the aqueous pKa. The obtained results have successfully been compared to literature ones obtained by other methods. It is concluded that the IS-CE method allows the determination of aqueous pKa values using low proportions of methanol, becoming then more accurate in the extrapolation procedure than other reference methods.

Ionizable Drug Self-Associations and the Solubility Dependence on pH: Detection of Aggregates in Saturated Solutions Using Mass Spectrometry (ESI-Q-TOF-MS/MS).

It is widely accepted that solubility-pH profiles of ionizable compounds follow the Henderson-Hasselbalch equation. However, several studies point out that compounds often undergo additional processes in saturated solutions, such as sub-micellar oligomerization, micellar aggregation, or drug-buffer complexation among others, which make the experimental profiles deviate from the behavior predicted by the Henderson-Hasselbalch equation. Often, the presence of additional processes is supported by the analysis of experimental data through solubility computer programs. However, the purpose of this work is to experimentally prove the aggregation phenomena for a series of bases for which deviations from the theoretical profile have been observed. To this end, five monoprotic bases (lidocaine, maprotiline, cyproheptadine, bupivacaine, and mifepristone) susceptible to form ionic aggregates in solution have been selected, and mass spectrometry has been the technique of choice to prove the presence of aggregation. High declustering potentials have been applied to prevent aggregates from forming in the ionization source of the mass spectrometer. In addition, haloperidol has been used as a negative control since according to its profile, it is not suspected to form ionic aggregates. In all instances, except for haloperidol, the analysis of the saturated solutions revealed the presence of mixed-charged dimers (aggregates formed by a neutral molecule and a charged one) and even trimers in the case of mifepristone and bupivacaine. For lidocaine, the most soluble of the compounds, the presence of neutral aggregates was also detected. These experiments support the hypothesis that the simple Henderson-Hasselbalch equation may explain the solubility-pH behavior of certain compounds, but it can be somewhat inaccurate in describing the behavior of many other substances.

Linear free energy relationship models for the retention of partially ionized acid-base compounds in reversed-phase liquid chromatography.

The LFER model of Abraham is applied to the retention of the neutral and ionic forms of 94 solutes in a C18 column and 40% v/v acetonitrile/water mobile phase. The results show that polarizability and cavity formation interactions increase retention, whereas dipole and hydrogen bonding interactions favours partition to the mobile phase and thus, they decrease retention. The coefficients of the ionic descriptors measure the effect of the electrostatic interactions and their contribution to partition of the cation or anion between the two mobile and stationary chromatographic phases. A new LFER model for application to the retention of partially dissociated acids and bases is derived averaging the descriptors of the neutral and ionic forms according to their degrees of ionization in the mobile phase. This new LFER model is satisfactorily compared to other literature modified Abraham models for a set of 498 retention data of partially dissociated acids and bases. All tested models require the calculation of the ionization degrees of the compounds at the measuring pH. Calculation of the ionization degrees in the chromatographic mobile phase (i.e. from pH and pKa in the eluent) give good correlations for all tested models. However, estimation of these ionization degrees from pH - pKa data in pure water gives biased estimations of the retention of the partially ionized solutes.

Synthesis and Characterization of a New Norfloxacin/Resorcinol Cocrystal with Enhanced Solubility and Dissolution Profile.

A new cocrystal of Norfloxacin, a poorly soluble fluoroquinolone antibiotic, has been synthetized by a solvent-mediated transformation experiment in toluene, using resorcinol as a coformer. The new cocrystal exists in both anhydrous and monohydrate forms with the same (1:1) Norfloxacin/resorcinol stoichiometry. The solubility of Norfloxacin and the hydrated cocrystal were determined by the shake-flask method. While Norfloxacin has a solubility of 0.32 ± 0.02 mg/mL, the cocrystal has a solubility of 2.64 ± 0.39 mg/mL, approximately 10-fold higher. The dissolution rate was tested at four biorelevant pH levels of the gastrointestinal tract: 2.0, 4.0, 5.5, and 7.4. In a first set of comparative tests, the dissolution rate of Norfloxacin and the cocrystal was determined separately at each pH value. Both solid forms showed the highest dissolution rate at pH 2.0, where Norfloxacin is totally protonated. Then, the dissolution rate decreases as pH increases. In a second set of experiments, the dissolution of the cocrystal was evaluated by a unique dissolution test, in which the pH dynamically changed from 2.0 to 7.4, stepping 30 min at each of the four biorelevant pH values. Results were quite different in this case, since dissolution at pH 2 affects the behavior of Norfloxacin at the rest of the pH values.

Determination of acidity constants at 37 °C through the internal standard capillary electrophoresis (IS-CE) method: internal standards and application to polyprotic drugs.

This work provides the pKa at the biorelevant temperature of 37 °C for a set of compounds proposed as internal standards for the internal standard capillary electrophoresis (IS-CE) method. This is a high throughput method that allows the determination of the acidity constants of compounds in a short time, avoiding the exact measurement of the pH of the buffers used. pH electrode calibration at 37 °C can be avoided too. In order to anchor the pKa values obtained through the IS-CE method in the pH scale, the acidity constant at 37 °C of some of the standards has been determined also by the reference potentiometric method. In general, a decrease in the pKa value is observed when changing the temperature from 25 to 37 °C, and the magnitude of the change depends on the nature of the compounds. Once the pKa values at 37 °C of the internal standards have been established, the method is applied to the determination of the acidity constants of seven polyprotic (5 diprotic and 2 triprotic) drugs. The obtained mobility-pH profiles show well-defined curves, and the fits provide precise pKa values. Due to the lack of reference data at 37 °C only the pKa values of labetalol can be compared to values from the literature, and a very good agreement is observed.

Optimization of experimental conditions for skin-PAMPA measurements.

In recent years, the parallel artificial membrane permeability assay (PAMPA) has been extended for prediction of skin permeation by developing an artificial membrane which mimics the stratum corneum structure, skin-PAMPA. In the present work, the different parameters affecting skin-PAMPA permeability, such as incubation time and stirring, have been studied to establish ideal assay conditions to generate quality data for a screening of active pharmaceutical ingredients (API) in early stage drug discovery. Another important parameter is membrane retention, which shows dependence on lipophilicity when compounds are in their neutral form. Furthermore, the stability of the membrane has been investigated at different pH values, especially at basic pHs. Finally, a good correlation between human skin permeability and skin-PAMPA permeability, with a large dataset (n = 46), has been established. The optimized assay conditions were an incubation time of 4 hours with stirring in a pH below 8. With all these considerations the thickness of the aqueous boundary layer is decreased as much as possible and the membrane stability is guaranteed.

Estimation of the octanol-water distribution coefficient of basic compounds by a cationic microemulsion electrokinetic chromatography system.

The octanol-water partition coefficient (Po/w), or the octanol-water distribution coefficient (Do/w) for ionized compounds, is a key parameter in the drug development process. In a previous work, this parameter was estimated through the retention factor measurements in a sodium dodecyl sulfate (SDS) - microemulsion electrokinetic chromatography (MEEKC) system for acidic compounds. Nonetheless, when ionized basic compounds were analyzed, undesirable ion pairs were formed with the anionic surfactant and avoided a good estimation of log Do/w. For this reason, an alternative MEEKC system based on a cationic surfactant has been evaluated to estimate Po/w or Do/w of neutral compounds and ionized bases. To this end, it has been characterized through the solvation parameter model (SPM) and compared to the octanol-water partition system. Results pointed out that both systems show a similar partition behavior. Hence, the log Po/w of a set of neutral compounds has been successfully correlated against the logarithm of the retention factor (log k) determined in this MEEKC system. Then, the log Do/w of 6 model bases have been estimated at different pH values and they have been compared to data from the literature, determined by the reference shake-flask and potentiometric methods. Good agreement has been observed between the literature and the estimated values when the base is neutral or partially ionized (up to 99% of ionization).

Estimation of the octanol-water distribution coefficient of acidic compounds by microemulsion electrokinetic chromatography.

The feasibility of extending the determination of the lipophilicity of partially ionized acids (log Do/w) by microemulsion electrokinetic chromatography (MEEKC) is tested. Theoretical considerations predict that a linear log Do/w vs. log k correlation can be obtained only when the neutral and ionic forms of an acid follow the same correlation equation and the slope of the correlation is unity. In practice, since the lipophilicity of the neutral acid is much higher than that of the ionic form and the correlation slope is not very different from 1, the general linear correlation for neutral compounds can be applied across most of the ionization range of the acid. The linear correlation between log Po/w and log k of 20 neutral solutes (calibration curve) has been established and extended to 6 acids used as models, tested across their full ionization range. log Do/w-pH, and log k-pH profiles have been obtained for these 6 acids, and plotted log Do/w against log k for any acid at any degree of ionization. Furthermore, the log Do/w of the acids has been estimated from the calibration curve and log k-pH profile, and compared to values in the literature determined using reference methods such as the shake-flask one. Accurate values have been obtained using the MEEKC method when the acids are in their neutral form or partially ionized (ionization degree, α < 0.995). However, this parameter is overestimated when the acids are highly or fully ionized (α ≈ 1). Finally, in order to test the applicability of this method, we have applied the same procedure to estimate log Do/w at pH = 7.4 (blood physiological pH) of a set of 30 additional compounds (including partially and fully ionized acids). The results at this pH follow the same trend observed in the 6 model acids, and validate the application of the method for Do/w determination, except when α is very close to 1.

Lecithin liposomes and microemulsions as new chromatographic phases.

Lecithins are phospholipidic mixtures that can be part of microemulsions and liposomes. In this work, ready-to-use preparations of lecithin have been tested as pseudostationary and mobile phases in EKC and LC, respectively. The selectivity of two EKC systems, one based on lecithin microemulsions (LMEEKC) and another on liposomes (LLEKC), and of a LC system based on lecithin microemulsions (MELC) has been evaluated through the solvation parameter model. In all cases, solute volume and hydrogen-bond basicity are the main descriptors that drive the partition process. While solute volume favors the retention of solutes, hydrogen-bond basicity has the contrary effect. In lecithin-based EKC systems the hydrogen-bond acidity of the solute leads to a higher retention while in the lecithin-based LC system a minor retention is produced. The three lecithin systems have been compared through the solvation parameter model to other chromatographic systems, most of them containing phospholipids. Principal component analysis reveals that lecithin systems cluster together with the other EKC systems based on phospholipids, with an immobilized artificial membrane (IAM) LC system, with the octanol/water reference partition system, and with a SDS-based microemulsion. Thus, they all show similar selectivity. However, the great advantage of using the ready-to use lecithin systems is that the laborious liposome preparation is avoided, and that their commercial availability makes them more affordable than IAM LC columns. Finally, taking into account that lecithin has a high semblance to the mammalian cell membranes composition, the ability of the three lecithin systems to mimic the pass of the solutes through the membranes has been evaluated. Experimental determinations have demonstrated that the skin partition of neutral solutes can be easily emulated, especially using the lecithin-microemulsion EKC method. The model is robust and shows good prediction ability.

Influence of the acid-base ionization of drugs in their retention in reversed-phase liquid chromatography.

The effect of the ionization in the RP-HPLC retention of 66 acid-base compounds, most of them drugs of pharmaceutical interest, is studied. The retention time of the compounds can be related to the pH measured in the mobile phase (pwsH) through the sigmoidal equations derived from distribution of the neutral and ionic forms of the drug into the stationary and mobile phases. Fitting of the obtained retention vs. pH profiles provides the retention times of the ionic and neutral forms and the pKa values of the drugs in the mobile phase (pwsKa). The obtained pwsKa values are linearly correlated to the pKa values in water (pwwKa) with two different correlations, one for neutral acids and another for neutral bases that reflect the different influence of the dielectric constant of the medium in ionization of acids and bases. The retention of the neutral species is well correlated to the octanol-water partition coefficient of the drugs as measure of the lipophilicity of the drug, which affects chromatographic retention. Also, the retention time of the ionized forms is related to the retention time of the neutral forms by two different linear correlations, one for anions and the other for cations. These last correlations point out the different retention behaviour of anions and cations: anions are less retained than cations of the same lipophilicity, as measured by the octanol-water partition coefficient of the neutral form. The different retention behaviour of anionic, cationic and neutral forms is confirmed by the hold-up times obtained from different approaches: pycnometry and retention times of anionic (KBr and KI) and neutral (DMSO) markers. Hold-up times obtained by pycnometric measurements agree with those obtained by retention of neutral markers (0.83-0.85 min), whereas hold-up time for anions is mobile phase pH dependent. At acidic pH it is similar to the hold-up time for neutral markers (0.83 min), but then it decreases with the increase of mobile phase pH to 0.65 min at pH 11. The decrease can be explained by the ionization of the silanols of the column and exclusion of anions by charge repulsion. Although not directly measured, the obtained retention data and correlations indicate hold-up time for cations are similar or slightly lower than hold-up time for neutral compounds (0.77-0.83 min). The model proposed and the correlations obtained can be very useful for its implementation in retention prediction algorithms for optimization of separation purposes.

Determination of the retention factor of ionizable compounds in microemulsion electrokinetic chromatography.

Determination of the retention factor of ionized compounds in microemulsion electrokinetic chromatography requires two mobility measurements at the same pH: one in the presence of the microemulsion and another in plain buffer. However, it has been observed that in some cases subtracting one mobility from another determined in a different medium leads to negative retention factors, which makes no sense from a chemical point of view. This indicates that there is some error in the process which has a direct impact when retention factors are used for further applications. Here, we evaluate how the components of the microemulsion confer different properties to the buffer medium, particularly varying the viscosity parameter (which is inversely related to mobility). Whereas sodium dodecyl sulfate, the surfactant used in the microemulsion, has little effect on the medium viscosity (only an increase of 5%-6%), the presence of 1-butanol, used as a stabilizer, increases it by around 30%. Meanwhile, heptane, which is used as an oil, provokes a slight decrease. Consequently, the mobilities obtained in the microemulsion system are shifted to higher values (less negative mobilities) compared to mobilities obtained in the aqueous buffer, and so one cannot be directly subtracted from the other. Since the microemulsion-buffer medium cannot be directly reproduced, we propose a correction that takes into account the variation of viscosities. This is determined from the electrophoretic mobility of the benzoate ion. As this ion does not interact with the microemulsion, the ratio of its mobilities (measured in plain buffer and microemulsion) is equivalent to the ratio of viscosities, and can be used as the correction factor for other measurements. Thus, mobilities in buffer and microemulsion media are placed on the same scale, overcoming the errors in retention factor determination.

Effect of vinylpyrrolidone polymers on the solubility and supersaturation of drugs; a study using the Cheqsol method.

The development of methods to increase the bioavailability of drugs is of great interest, especially for those which are poorly soluble or permeable. One of the strategies to enhance the solubility (which in turn has the potential of increase bioavailability) of drugs is the use of additives in the formulation process, so that the drug can stay supersaturated in biological fluids for a period of time long enough to allow absorption. The use of polymers as pharmaceutical excipients in order to stabilize the supersaturation of drugs is common practice. In this work, the ability of different polymers of vinylpyrrolidone (K-12, K-17, K-25, K-29/32, K-90) and a copolymer of vinylpyrrolidone and vinylacetate (S-630) have been tested for their impact on the supersaturation of drugs. Sixteen drugs of different chemical nature have been selected, and analyzed using the Cheqsol method. The results of the drug alone, and of physical mixtures with the different polymers at several polymer:drug ratios have been compared in terms of supersaturation extent and duration. It has been observed that acidic compounds displayed enhanced solubility in different ways: sometimes the supersaturated state of the drug is maintained for a long time, due to the precipitation of an amorphous solid, as determined by X-ray diffraction studies; on other occasions supersaturation increases but only for a short time, compared to the drug alone, and then the drug precipitates to a crystalline form. Only a few basic drugs displayed enhanced solubility in the presence of PVP polymers, in contrast to acidic compounds.

Molecular interactions between warfarin and human (HSA) or bovine (BSA) serum albumin evaluated by isothermal titration calorimetry (ITC), fluorescence spectrometry (FS) and frontal analysis capillary electrophoresis (FA/CE).

Interaction thermodynamics between warfarin, a very popular anticoagulant, and Sudlow I binding site of human (HSA) or bovine (BSA) serum albumin have been examined in strictly controlled experimental conditions (HEPES buffer 50 mM, pH 7.4 and 25 °C) by means of isothermal titration calorimetry (ITC), fluorescence spectrometry (FS) and frontal analysis capillary electrophoresis (FA/CE). Each technique is based on measurements of a different property of the biochemical system, and then the results allow a critical discussion about the suitability of each approach to estimate the drug-protein binding parameters. The strongest interaction step is properly evaluated by the three assayed approaches being the derived binding constants strongly consistent: from 4 × 104 to 7 × 104 for HSA and from 0.8 × 105 to 1.2 × 105 for BSA. Binding enthalpy variations also show consistent results: -5.4 and -5.6 Kcal/mol for HSA and -4.3 and -3.7 Kcal/mol for BSA, as measured by ITC and FS, respectively. Further high order interaction events for both albumins are detected only by FA/CE.

A high-fat high-sucrose diet affects the long-term metabolic fate of grape proanthocyanidins in rats.

PURPOSE: Polyphenol metabolites are key mediators of the biological activities of polyphenols. This study aimed to evaluate the long-term effects of a high-fat high-sucrose (HFHS) diet on the metabolism of proanthocyanidins from grape seed extract (GSE). METHODS: Adult female Wistar-Kyoto rats were fed a standard (STD) or HFHS diet supplemented or not with GSE for 16 weeks. PA metabolites were determined by targeted HPLC-MS/MS analysis. RESULTS: A lower concentration of total microbial-derived PA metabolites was present in urine and the aqueous fraction of faeces in the HFHS + GSE group than in the STD + GSE group. In contrast, a tendency towards the formation of conjugated (epi)catechin metabolites in the HFHS + GSE group was observed. CONCLUSIONS: These results show that a HFHS diet significantly modifies PA metabolism, probably via: (1) a shift in microbial communities not counteracted by the polyphenols themselves; and (2) an up-regulation of hepatic enzymes.

Influence of omega-3 PUFAs on the metabolism of proanthocyanidins in rats.

Studies of the bioavailability of proanthocyanidins usually consider them independently of other dietary constituents, while there is a tendency in the field of functional foods towards the combination of different bioactive compounds in a single product. This study examined the long-term effects of ω-3 polyunsaturated fatty acids of marine origin on the metabolic fate of grape proanthocyanidins. For this, female adult Wistar-Kyoto rats were fed (18weeks) with a standard diet supplemented or not with eicosapentaenoic acid/docosahexaenoic acid (1:1, 16.6g/kg feed), proanthocyanidin-rich grape seed extract (0.8g/kg feed) or both. A total of 39 microbial-derived metabolites and 16 conjugated metabolites were detected by HPLC-MS/MS either in urine or in the aqueous fraction of feces. An unexpected significant increase in many proanthocyanidin metabolites in urine and feces was observed in the group supplemented with ω-3 polyunsaturated fatty acids group as compared to the animals fed a standard diet, which contains a small amount of polyphenols. However, proanthocyanidin metabolites in rats given ω-3 polyunsaturated fatty acids and grape seed extract did not significantly differ from those in the group supplemented only with grape seed extract. It was concluded that ω-3 polyunsaturated fatty acids collaborate in the metabolism of polyphenols when present at low doses in the feed matrix, while the capacity of ω-3 polyunsaturated fatty acids to induce microbiota transformations when proanthocyanidins are present at high doses is not relevant compared to that of polyphenols themselves.

Modeling Aquatic Toxicity through Chromatographic Systems.

Environmental risk assessment requires information about the toxicity of the growing number of chemical products coming from different origins that can contaminate water and become toxicants to aquatic species or other living beings via the trophic chain. Direct toxicity measurements using sensitive aquatic species can be carried out but they may become expensive and ethically questionable. Literature refers to the use of chromatographic measurements that correlate to the toxic effect of a compound over a specific aquatic species as an alternative to get toxicity information. In this work, we have studied the similarity in the response of the toxicity to different species and we have selected eight representative aquatic species (including tadpoles, fish, water fleas, protozoan, and bacteria) with known nonspecific toxicity to chemical substances. Next, we have selected four chromatographic systems offering good perspectives for surrogation of the eight selected aquatic systems, and thus prediction of toxicity from the chromatographic measurement. Then toxicity has been correlated to the chromatographic retention factor. Satisfactory correlation results have been obtained to emulate toxicity in five of the selected aquatic species through some of the chromatographic systems. Other aquatic species with similar characteristics to these five representative ones could also be emulated by using the same chromatographic systems. The final aim of this study is to model chemical products toxicity to aquatic species by means of chromatographic systems to reduce in vivo testing.