Toxicity of Bromo-DragonFLY as a New Psychoactive Substance: Application of In Silico Methods for the Prediction of Key Toxicological Parameters Important to Clinical and Forensic Toxicology.

Bromo-DragonFLY is a synthetic new psychoactive substance (NPS) that has gained attention due to its powerful and long-lasting hallucinogenic effects, legal status, and widespread availability. This study aimed to use various in silico toxicology methods to predict key toxicological parameters for Bromo-DragonFLY, including acute toxicity (LD50), genotoxicity, cardiotoxicity, health effects, and the potential for endocrine disruption. The results indicate significant acute toxicity with noticeable variations across different species, a low likelihood of genotoxic potential suggesting potential DNA damage, and a notable risk of cardiotoxicity associated with inhibition of the hERG channel. Evaluation of endocrine disruption suggests a low probability of Bromo-DragonFLY interacting with the estrogen receptor α (ER-α), indicating minimal estrogenic activity. These insights from in silico investigations are important for advancing our understanding of this NPS in forensic and clinical toxicology. These initial toxicological examinations establish a foundation for future research efforts and contribute to developing risk assessment and management strategies for using and misusing NPS.

The prediction of acute toxicity (LD50) for organophosphorus-based chemical warfare agents (V-series) using toxicology in silico methods.

Nerve agents are organophosphate chemical warfare agents that exert their toxic effects by irreversibly inhibiting acetylcholinesterase, affecting the breakdown of the neurotransmitter acetylcholine in the synaptic cleft. Due to the risk of exposure to dangerous nerve agents and for animal welfare reasons, in silico methods have been used to assess acute toxicity safely. The next-generation risk assessment (NGRA) is a new approach for predicting toxicological parameters that can meet modern requirements for toxicological research. The present study explains the acute toxicity of the examined V-series nerve agents (n = 9) using QSAR models. Toxicity Estimation Software Tool (ver. 4.2.1 and ver. 5.1.2), QSAR Toolbox (ver. 4.6), and ProTox-II browser application were used to predict the median lethal dose. The Simplified Molecular Input Line Entry Specification (SMILES) was the input data source. The results indicate that the most deadly V-agents were VX and VM, followed by structural VX analogues: RVX and CVX. The least toxic turned out to be V-sub x and Substance 100A. In silico methods for predicting various parameters are crucial for filling data gaps ahead of experimental research and preparing for the upcoming use of nerve agents.

The estimation of acute oral toxicity (LD50) of G-series organophosphorus-based chemical warfare agents using quantitative and qualitative toxicology in silico methods.

The idea of this study was the estimation of the theoretical acute toxicity (t-LD50, rat, oral dose) of organophosphorus-based chemical warfare agents from the G-series (n = 12) using different in silico methods. Initially identified in Germany, the G-type nerve agents include potent compounds such as tabun, sarin, and soman. Despite their historical significance, there is a noticeable gap in acute toxicity data for these agents. This study employs qualitative (STopTox and AdmetSAR) and quantitative (TEST; CATMoS; ProTox-II and QSAR Toolbox) in silico methods to predict LD50 values, offering an ethical alternative to animal testing. Additionally, we conducted quantitative extrapolation from animals, and the results of qualitative tests confirmed the acute toxicity potential of these substances and enabled the identification of toxicophoric groups. According to our estimations, the most lethal agents within this category were GV, soman (GD), sarin (GB), thiosarin (GBS), and chlorosarin (GC), with t-LD50 values (oral administration, extrapolated from rat to human) of 0.05 mg/kg bw, 0.08 mg/kg bw, 0.12 mg/kg bw, 0.15 mg/kg bw, and 0.17 mg/kg bw, respectively. On the contrary, compounds with a cycloalkane attached to the phospho-oxygen linkage, specifically methyl cyclosarin and cyclosarin, were found to be the least toxic, with values of 2.28 mg/kg bw and 3.03 mg/kg bw. The findings aim to fill the knowledge gap regarding the acute toxicity of these agents, highlighting the need for modern toxicological methods that align with ethical considerations, next-generation risk assessment (NGRA) and the 3Rs (replacement, reduction and refinement) principles.

The acute toxicity of Novichok's degradation products using quantitative and qualitative toxicology in silico methods.

The emergence of Novichok agents, potent organophosphorus nerve agents, has spurred the demand for advanced analytical methods and toxicity assessments as a result of their involvement in high-profile incidents. This study focuses on the degradation products of Novichok agents, particularly their potential toxic effects on biological systems. Traditional in vivo methods for toxicity evaluation face ethical and practical constraints, prompting a shift toward in silico toxicology research. In this context, we conducted a comprehensive qualitative and quantitative analysis of acute oral toxicity (AOT) for Novichok degradation products, using various in silico methods, including TEST, CATMoS, ProTox-II, ADMETlab, ACD/Labs Percepta, and QSAR Toolbox. Adopting these methodologies aligns with the 3Rs principle, emphasising Replacement, Reduction, and Refinement in the realm of toxicological studies. Qualitative assessments with STopTox and admetSAR revealed toxic profiles for all degradation products, with predicted toxicophores highlighting structural features responsible for toxicity. Quantitative predictions yielded varied estimates of acute oral toxicity, with the most toxic degradation products being EOPAA, MOPGA, MOPAA, MPGA, EOPGA, and MPAA, respectively. Structural modifications common to all examined hydrolytic degradation products involve substituting the fluorine atom with a hydroxyl group, imparting consequential effects on toxicity. The need for sophisticated analytical techniques for identifying and quantifying Novichok degradation products is underscored due to their inherent reactivity. This study represents a crucial step in unravelling the complexities of Novichok toxicity, highlighting the ongoing need for research into its degradation processes to refine analytical methodologies and fortify readiness against potential threats.

The prediction of hydrolysis and biodegradation of organophosphorus-based chemical warfare agents (G-series and V-series) using toxicology in silico methods.

Nerve agents (G- and V-series) are a group of extremely toxic organophosphorus chemical warfare agents that we have had the opportunity to encounter many times on a massive scale (Matsumoto City, Tokyo subway and Gulf War). The threat of using nerve agents in terrorist attacks or military operations is still present, even with establishing the Chemical Weapons Convention as the legal framework. Understanding their environmental sustainability and health risks is critical to social security. Due to the risk of contact with dangerous nerve agents and animal welfare considerations, in silico methods were used to assess hydrolysis and biodegradation safely. The environmental fate of the examined nerve agents was elucidated using QSAR models. The results indicate that the investigated compounds released into the environment hydrolyse at a different rate, from extremely fast (<1 day) to very slow (over a year); V-agents undergo slower hydrolysis compared to G-agents. V-agents turned out to be relatively challenging to biodegrade, the ultimate biodegradation time frame of which was predicted as weeks to months, while for G-agents, the overwhelming majority was classified as weeks. In silico methods for predicting various parameters are critical to preparing for the forthcoming application of nerve agents.

Toxicity of the New Psychoactive Substance (NPS) Clephedrone (4-Chloromethcathinone, 4-CMC): Prediction of Toxicity Using In Silico Methods for Clinical and Forensic Purposes.

This study reports the first application of in silico methods to assess the toxicity of 4-chloromethcathinone (4-CMC), a novel psychoactive substance (NPS). Employing advanced toxicology in silico tools, it was possible to predict crucial aspects of the toxicological profile of 4-CMC, including acute toxicity (LD50), genotoxicity, cardiotoxicity, and its potential for endocrine disruption. The obtained results indicate significant acute toxicity with species-specific variability, moderate genotoxic potential suggesting the risk of DNA damage, and a notable cardiotoxicity risk associated with hERG channel inhibition. Endocrine disruption assessment revealed a low probability of 4-CMC interacting with estrogen receptor alpha (ER-α), suggesting minimal estrogenic activity. These insights, derived from in silico studies, are critical in advancing the understanding of 4-CMC properties in forensic and clinical toxicology. These initial toxicological findings provide a foundation for future research and aid in the formulation of risk assessment and management strategies in the context of the use and abuse of NPSs.

Multimodal Imaging Using Raman Spectroscopy and FTIR in a Single Analytical Instrument with a Microscope (Infrared Raman Microscopy AIRsight, Shimadzu): Opportunities and Applications.

Raman spectroscopy and Fourier transform infrared (FTIR) spectroscopy are powerful analytical techniques widely used separately in different fields of study. Integrating these two powerful spectroscopic techniques into one device represents a groundbreaking advance in multimodal imaging. This new combination which merges the molecular vibrational information from Raman spectroscopy with the ability of FTIR to study polar bonds, creates a unique and complete analytical tool. Through a detailed examination of the microscope's operation and case studies, this article illustrates how this integrated analytical instrument can provide more thorough and accurate analysis than traditional methods, potentially revolutionising analytical sample characterisation. This article aims to present the features and possible uses of a unified instrument merging FTIR and Raman spectroscopy for multimodal imaging. It particularly focuses on the technological progress and collaborative benefits of these two spectroscopic techniques within the microscope system. By emphasising this approach's unique benefits and improved analytical capabilities, the authors aim to illustrate its applicability in diverse scientific and industrial sectors.

Gold Nanoparticles (AuNPs)-Toxicity, Safety and Green Synthesis: A Critical Review.

In recent years, the extensive exploration of Gold Nanoparticles (AuNPs) has captivated the scientific community due to their versatile applications across various industries. With sizes typically ranging from 1 to 100 nm, AuNPs have emerged as promising entities for innovative technologies. This article comprehensively reviews recent advancements in AuNPs research, encompassing synthesis methodologies, diverse applications, and crucial insights into their toxicological profiles. Synthesis techniques for AuNPs span physical, chemical, and biological routes, focusing on eco-friendly "green synthesis" approaches. A critical examination of physical and chemical methods reveals their limitations, including high costs and the potential toxicity associated with using chemicals. Moreover, this article investigates the biosafety implications of AuNPs, shedding light on their potential toxic effects on cellular, tissue, and organ levels. By synthesizing key findings, this review underscores the pressing need for a thorough understanding of AuNPs toxicities, providing essential insights for safety assessment and advancing green toxicology principles.

What do we currently know about Novichoks? The state of the art.

Novichok is the name given to the group of nerve agents created stealthily in the later phases of the Cold War by the Soviet Union. Constitute the fourth generation of chemical warfare agents; like other nerve agents, they are organophosphorus compounds designed to be incurable and undetectable. The mechanism of action is based on the non-competitive and irreversible inhibition of acetylcholinesterase. Due to their enormous toxicity, Novichoks have become attractive targets for terrorists. However, little information is known about the identity of nerve agents. Furthermore, these compounds have never been submitted to the Chemical Weapons Convention. Our article aspires to provide a general overview of Novichoks knowledge. As part of this, we reviewed the available literature data to answer the question, what are Novichoks? In addition to the physical and chemical properties of A-agents, synthesis, mechanism of action, and toxicity of nerve agents were also reviewed. We hope that this review will highlight the tremendous threat posed by nerve agents and will inspire further studies on the interdisciplinary aspects of these compounds.

Application of toxicology in silico methods for prediction of acute toxicity (LD50) for Novichoks.

Novichoks represent the fourth generation of chemical warfare agents with paralytic and convulsive effects, produced clandestinely during the Cold War by the Soviet Union. This novel class of organophosphate compounds is characterised by severe toxicity, which, for example, we have already experienced three times (Salisbury, Amesbury, and Navalny's case) as a society. Then the public debate about the true nature of Novichoks began, realising the importance of examining the properties, especially the toxicological aspects of these compounds. The updated Chemical Warfare Agents list registers over 10,000 compounds as candidate structures for Novichoks. Consequently, conducting experimental research for each of them would be a huge challenge. Additionally, due to the enormous risk of contact with hazardous Novichoks, in silico assessments were applied to estimate their toxicity safely. In silico toxicology provides a means of identifying hazards of compounds before synthesis, helping to fill gaps and guide risk minimisation strategies. A new approach to toxicology testing first considers the prediction of toxicological parameters, eliminating unnecessary animal studies. This new generation risk assessment (NGRA) can meet the modern requirements of toxicological research. The present study explains, using QSAR models, the acute toxicity of the Novichoks studied (n = 17). The results indicate that the toxicity of Novichoks varies. The deadliest turned out to be A-232, followed by A-230 and A-234. On the other hand, the "Iranian" Novichok and C01-A038 compounds turned out to be the least toxic. Developing reliable in silico methods to predict various parameters is essential to prepare for the upcoming use of Novichoks.

Toxicological Aspects, Safety Assessment, and Green Toxicology of Silver Nanoparticles (AgNPs)-Critical Review: State of the Art.

In recent years, research on silver nanoparticles (AgNPs) has attracted considerable interest among scientists because of, among other things, their alternative application to well-known medical agents with antibacterial properties. The size of the silver nanoparticles ranges from 1 to 100 nm. In this paper, we review the progress of research on AgNPs with respect to the synthesis, applications, and toxicological safety of AgNPs, and the issue of in vivo and in vitro research on silver nanoparticles. AgNPs' synthesis methods include physical, chemical, and biological routes, as well as "green synthesis". The content of this article covers issues related to the disadvantages of physical and chemical methods, which are expensive and can also have toxicity. This review pays special attention to AgNP biosafety concerns, such as potential toxicity to cells, tissues, and organs.

The Critical Assessment of Oxidative Stress Parameters as Potential Biomarkers of Carbon Monoxide Poisoning.

In conventional clinical toxicology practice, the blood level of carboxyhemoglobin is a biomarker of carbon monoxide (CO) poisoning but does not correspond to the complete clinical picture and the severity of the poisoning. Taking into account articles suggesting the relationship between oxidative stress parameters and CO poisoning, it seems reasonable to consider this topic more broadly, including experimental biochemical data (oxidative stress parameters) and patients poisoned with CO. This article aimed to critically assess oxidative-stress-related parameters as potential biomarkers to evaluate the severity of CO poisoning and their possible role in the decision to treat. The critically set parameters were antioxidative, including catalase, 2,2-diphenyl-1-picryl-hydrazyl, glutathione, thiol and carbonyl groups. Our preliminary studies involved patients (n = 82) admitted to the Toxicology Clinical Department of the University Hospital of Jagiellonian University Medical College (Kraków, Poland) during 2015-2020. The poisoning was diagnosed based on medical history, clinical symptoms, and carboxyhemoglobin blood level. Blood samples for carboxyhemoglobin and antioxidative parameters were collected immediately after admission to the emergency department. To evaluate the severity of the poisoning, the Pach scale was applied. The final analysis included a significant decrease in catalase activity and a reduction in glutathione level in all poisoned patients based on the severity of the Pach scale: I°-III° compared to the control group. It follows from the experimental data that the poisoned patients had a significant increase in level due to thiol groups and the 2,2-diphenyl-1-picryl-hydrazyl radical, with no significant differences according to the severity of poisoning. The catalase-to-glutathione and thiol-to-glutathione ratios showed the most important differences between the poisoned patients and the control group, with a significant increase in the poisoned group. The ratios did not differentiate the severity of the poisoning. The carbonyl level was highest in the control group compared to the poisoned group but was not statistically significant. Our critical assessment shows that using oxidative-stress-related parameters to evaluate the severity of CO poisoning, the outcome, and treatment options is challenging.

The Development of New Methodology for Determination of Vincristine (VCR) in Human Serum Using LC-MS/MS-Based Method for Medical Diagnostics.

In this article, we have presented the development and validation of a rapid and sensitive reversed-phase liquid chromatography with tandem mass spectrometry (LC-MS/MS) method for the determination of vincristine (VCR) in patient serum samples. Chromatographic separation was achieved on a Kinetex® (Singapore) column using a mobile phase consisting of 25 mM acetic acid and 0.3% formic acid (A) and methanol (B) in a gradient elution mode at a flow rate of 0.3 mL/min. The VCR and internal standard (vinblastine) were monitored using the multiple reaction monitoring mode under positive electrospray ionization. The lower limit of quantification (LLOQ) was 0.67 ng/mL, and the upper limit of quantification (ULOQ) was 250 ng/mL for VCR. The calculated values of LOD and LOQ for VCR were 0.075 and 0.228 ng/mL, respectively. The calibration curve was linear over the VCR concentration range of 1.0−250 ng/mL in serum. The intra- and inter-day precision and precision were within the generally accepted criteria for the bioanalytical method (<15%). The method was successfully applied to the analysis of serum samples in clinical practice.

Ni and Cr impurities profile in Valeriana officinalis L., radix-based herbal medicinal product available in polish pharmacies due to ICH Q3D guideline.

Elemental impurities (EIs) profiling in final pharmaceutical products is often not adequately treated, however it is crucial problem in pharmaceutical analysis by reason of the various regulatory authorities (like ICH Q3D guideline). EIs in pharmaceuticals may arise from numerous sources of which the herbal ingredients are not a frequent subject of pharmaceutical analyses. However, based on number of traditional use registrations per year for herbal medicinal products (HMPs) in the EU, it can be stated that monocomponent HMPs are still very popular for use. Due to the high frequency of use, exposure to EIs from HMPs may be high during long-term use. The aim of our article was Ni and Cr impurities profiling of Valeriana officinalis L., radix (Valerian root) as an example of the HMP available in Polish pharmacies for the relief of mild nervous tension and sleep disorders. The choice of metals was justified by: (1) a single dose of Ni administered via oral route can induce dermatitis in nickel-sensitised individuals; (2) Cr is a very problematic element from toxicological point of view. Our results indicate that the standards of the ICH Q3D guideline are met for all EIs.

Biological consequences of zinc deficiency in the pathomechanisms of selected diseases.

From many points of view, zinc is one of the most important trace elements in biological systems. Many articles describe the well-known role of this metal in human physiology and pathophysiology, but in the related literature, there is a lack of current and reliable reviews of the role of zinc deficiency in many diseases. In this article, we describe the role of zinc deficiency in the oxidative stress control, immune response, proliferation, and pathogenesis and pathophysiology of selected diseases such as depression, cardiovascular diseases, diabetes mellitus, Alzheimer's disease, and Wilson's disease.

The toxicological analysis of lead impurities in traditional herbal medicinal product with Thymi herba (Thymus vulgaris L. and Thymus zygis L.) from pharmacies in Poland: the impurity profile and comprehensive toxicological risk assessment.

Traditional herbal medicinal products (THMP) with Thymi herba (Thymus vulgaris L. and Thymus zygis L.) are very popular among the European population. The idea of our study was the toxicological analysis of lead impurities in THMP with Thymi herba obtained from Polish pharmacies. For this purpose, we prepared the impurity profiles and comprehensive toxicological risk assessment. The Pb impurity profiles show lead impurities in all samples investigated (in the range: 2.15-6.99 µg/L). The estimations of Pb impurities in the single dose (32.25-105.01 ng/single dose) and in the daily dose (64.50-210.00 ng/day) were based on the posology recommended by the manufacturers. All the results obtained meet the standards of the ICH Q3D (R1) guideline on elemental impurities according to the levels of Pb. It can be concluded that all investigated THMPs with Thymi herba available in Poland should not represent any health risk to adults.

The comprehensive prediction of carcinogenic potency and tumorigenic dose (TD50) for two problematic N-nitrosamines in food: NMAMPA and NMAMBA using toxicology in silico methods.

The identification and toxicological assessment of potential carcinogens is of paramount importance for public health and safety. This study aimed to predict the carcinogenic potency and tumorigenic dose (TD50) for two problematic N-nitrosamines (N-NAs) commonly found in food: N-2-methylpropyl-N-1-methylacetonylnitrosamine (NMAMPA, CAS: 93755-83-0) and N-3-Methylbutyl-N-1-methylacetonylnitrosamine (NMAMBA, CAS: 71016-15-4). To achieve this goal, in silico toxicology methods were employed due to their practical applications and potential in risk assessment. The justification for conducting these studies was incoherent results published by the European Food Safety Authority (EFSA). For this purpose, we applied various in silico tools, including qualitative methods (ToxTree, ProTox II and CarcinoPred-EL) and quantitative methods (QSAR Toolbox and LAZAR) were applied to predict the carcinogenic potency. These tools leverage computational approaches to analyze chemical structures for finding toxicophores and generating predictions, making them efficient alternatives to traditional in vivo experiments. The results obtained indicated that N-NAs are carcinogenic compounds, and quantitative data was obtained in the form of estimated doses of TD50 for the compounds tested.

Toxicity of minoxidil - Comprehensive in silico prediction of main toxicity endpoints: Acute toxicity, irritation of skin and eye, genetic toxicity, health effect, cardiotoxicity and endocrine system disruption.

This article focusses on elucidating the toxicological profile of minoxidil, a widely used pharmacological agent for alopecia, through the application of in silico methods (Percepta ACD/Labs software). This research is driven by the need to understand key toxicological endpoints: acute toxicity, skin and eye irritation, genetic toxicity, cardiotoxicity, disruption of the endocrine system, and estimation of various health effects due to the lack of experimental data for this drug. These parameters are critically evaluated to meet the stringent requirements of the pharmaceutical industry's safety assessments. The results obtained for acute toxicity (LD50 for rats and mouse) indicate that minoxidil exhibits a species-dependent acute toxicity profile e.g. 51 mg/kg bw for intravenous administration in mice. The predicted health effects indicate a 93% risk to the gastrointestinal system, 54% for the kidneys, 52% for the liver, 42% for the blood and lungs, and 39% for the cardiovascular system. The prediction of genotoxicity suggests a moderate probability (48%) of inducing a positive Ames test result. Furthermore, moderate inhibition of the hERG channel indicates potential cardiac risks of Minoxidil. Based on the information obtained, we propose subjecting minoxidil to additional toxicological assessments. The successful adoption of these in silico methodologies aligns with the 3 R s principle (replacement, reduction, and refinement) in the field of modern toxicological studies of minoxidil, all without the use of laboratory animals for the novelty of our toxicity assessment.

Hazardous elements in plastic and rubber granules as infill material from sports facilities? Field Portable-XRF spectroscopy as 'white analytical technique' reveals hazardous elements in fall sports facilities in Rzeszów (Podkarpackie, Poland).

Plastic and rubber granules are commonly used as infill material in all-weather sports facilities, providing an ideal activity surface for millions of Europeans on a daily basis. However, concerns have been raised about the presence of hazardous elements in these granules, which can pose risks both to the environment and human health. Our study focusses on the elemental composition of rubber granules used in fall sports facilities in Rzeszów, (Podkarpackie, Poland) using field portable X-ray fluorescence (FP-XRF) as a non-destructive and 'white analytical technique'. The results show the content of Zn, Fe, Cr, Ba, Br, Ti, Cu, Cd, As, Au, Bi, Pb, Ni, Sb, and Sn in the rubber granule samples. This study highlights the need for stringent quality control measures and regulations to ensure the safety of all-weather sports facilities and protect the well-being of sportsman. When modern FP-XRF spectrometry is employed as a "white analytical technique," for the first time it becomes possible to identify the presence of hazardous elements, addressing the pressing concerns highlighted by the ECHA and enabling proactive measures to mitigate potential risks. This approach ensures the protection of the health and sustainability of sports facilities, contributing to the ongoing hot topics in the field.

The comprehensive strategy in the human health risk assessment of total chromium impurities in cough syrups with Marshmallow Root (Althaea officinalis) available in Polish pharmacies: regulatory aspects and special emphasis on Cr(VI) mode of action.

Chromium, which can currently only be considered pharmacologically active and not an essential element, is a very intriguing elemental impurity in final pharmaceutical products, especially traditional herbal medicinal products. This kind of traditional herbal medicinal product with Marshmallow root (Althaea officinalis L., radix) registered in the EU is widely used among the European population. The aim of this article is to propose a double regulatory strategy in assessing the human health risk of total chromium impurities in cough syrups with Marshmallow Root (Althaea officinalis) available in Polish pharmacies. We applied the strategy based on the requirements of the ICH Q3D (R1) guideline for the assessment of Cr impurities in final traditional herbal medicinal products with Marshmallow Root registered in the EU. Furthermore, we applied the strategy based on the concept of margin of exposure (MoE) considering Cr(VI) genotoxicity mode of action (MOA) and based on BMD10 for Cr(VI) as a point of departure (PoD). The total Cr content was in the range: 1.12-9.61 µg/L (in comparison with the ICH Q3D R1 guidelines: 1100 µg/g). Total Cr levels in a single dose were relatively high compared to raw results, but were not a threat to patients. Comparison of estimated results with oral PDE value for Cr in final drugs suggested by the ICH Q3D R1 guideline (10,700 µg/day) show that all the products analyzed were below this value (the highest result was 278.40 ng/day). Despite conservative assumptions, the MoE values obtained for Cr in daily dose for each Marshmallow Root cough syrup were above 10,000; therefore, exposure to Cr would not cause a health risk for specific population groups (children and adults). It can be summarized that each of the phytopharmaceuticals analysed with Marshmallow root available in Polish pharmacies does not represent a health hazard to patients. We confirm the safety of Cr impurities by applying a double regulatory strategy without the application of an expansive and demanding HPLC-ICP-MS technique for Cr speciation.