Strategies for quantifying C(60) fullerenes in environmental and biological samples and implications for studies in environmental health and ecotoxicology

Fullerenes are sphere-like molecules with unique physico-chemical properties, which render them of particular interest in biomedical research, consumer products and industrial applications. Human and environmental exposure to fullerenes is not a new phenomenon, due to a long history of hydrocarbon-combustion sources, and will only increase in the future, as incorporation of fullerenes into consumer products becomes more widespread for use as anti-aging, anti-bacterial or anti-apoptotic agents.An essential step in the determination of biological effects of fullerenes (and their surface-functionalized derivatives) is establishment of exposure-assessment techniques. However, in ecotoxicological studies, quantification of fullerenes is performed infrequently because robust, uniformly applicable analytical approaches have yet to be identified, due to the wide variety of sample types. Moreover, the unique physico-chemistry of fullerenes in aqueous matrices requires reassessment of conventional analytical approaches, especially in more complex biological matrices (e.g., urine, blood, plasma, milk, and tissue).Here, we present a review of current analytical approaches for the quantification of fullerenes and propose a consensus approach for determination of these nanomaterials in a variety of environmental and biological matrices.     

Electrochemistry of Fullerenes and Their Derivatives

Applications of voltammetry and electrosynthesis in the chemistry of fullerenes and their derivatives are considered. Effect of structural factors and environment on the reduction potentials of fullerenes and stability of their anion intermediates are analyzed. Emphasis is laid on the reduction of phosphorylated methanofullerenes.     

Liquid chromatography–mass spectrometry for C60 fullerene analysis: optimisation and comparison of three ionisation techniques

The increasing use and production of nanomaterials have led to growing concern over the release of new pollutants to the environment. Fullerenes have been a subject of intense research, both because of their unique chemistry and because of technological applications. The development of analytical methods to quantify the fullerenes in complex sample matrices is a crucial step in the study of their occurrence and exposure, and thus in risk assessment. This paper reports the development and optimisation of a method combining liquid chromatography with ion-trap mass spectrometry (LC-ITMS) for analysis of the fullerene C(60). Under the optimised chromatogram conditions, a C(18) analytical column had good selectivity for fullerenes C(60) and C(70), with retention times of 3.0 and 4.1 min, respectively. Mass spectrometric detection was tested and optimised using three common ionisation techniques-atmospheric-pressure chemical ionisation (APCI), atmospheric-pressure photoionisation (APPI), and electrospray ionisation (ESI). The molecular ion was most abundant for C (60) (-) (m/z=720) in APCI and APPI, whereas adduct ions were formed with the molecular ion in ESI. Finally, the performance of the three ionisation techniques examined was compared by use of five validation criteria. The instrument detection limit (8 ng mL(-1)), quantification limit (27 ng mL(-1)), detection sensitivity (90.2 ng mL(-1)), linear range (8-1,000 ng mL(-1)), and repeatability (15 %) of APPI make it the most promising ionisation technique for fullerene C(60) analysis.     

Characterisation and determination of fullerenes: A critical review

A prominent sector of nanotechnology is occupied by a class of carbon-based nanoparticles known as fullerenes. Fullerene particle size and shape impact in how easily these particles are transported into and throughout the environment and living tissues. Currently, there is a lack of adequate methodology for their size and shape characterisation, identification and quantitative detection in environmental and biological samples. The most commonly used methods for their size measurements (aggregation, size distribution, shape, etc.), the effect of sampling and sample treatment on these characteristics and the analytical methods proposed for their determination in complex matrices are discussed in this review. For the characterisation and analysis of fullerenes in real samples, different analytical techniques including microscopy, spectroscopy, flow field-flow fractionation, electrophoresis, light scattering, liquid chromatography and mass spectrometry have been reported. The existing limitations and knowledge gaps in the use of these techniques are discussed and the necessity to hyphenate complementary ones for the accurate characterisation, identification and quantitation of these nanoparticles is highlighted.     

Low-Cost Synthesis of Fullerene Derivatives

Refined mixed fullerenes were used as a reagent in known organic reactions instead of the pure fullerene C₆₀ with aim to find an alternative, low-cost method for the synthesis of fullerene derivatives potentially exhibiting photoconductive properties. The isolation of C₆₀ or C₇₀ in clean form without admixtures requires the use of large quantities of toluene or other nonpolar solvents, polluting the environment and multiplying the production cost. 1,3-Dipolar cycloaddition of azomethine ylide to fullerite was chosen because this reaction is one of the most widely used for fullerene functionalization, producing material possibly presenting photoinducing behavior. The data showed that the use of the cheaper mixed fullerenes instead of pure C₆₀ leads to the isolation of the same expected products with similar yields. The photoelectric properties of mixed fullerenes and their organic derivatives were also examined. A slightly semiconductive behavior was confirmed as well as a noticeable photoresponse.

Supplemental materials are available for this article. Go to the publisher's online edition of Synthetic Communications® to view the free supplemental file.     

Occurrence of pharmaceutical metabolites and transformation products in the aquatic environment of the Mediterranean area

The presence of pharmaceutically active substances (PhACs) in aquatic ecosystems is of great concern due to their constant occurrence in different water systems and potential negative effects on the quality of water and living organisms. After consumption, PhACs are excreted as the parent compound, and/or as free or conjugated metabolites, which might finally arrive to surface and ground waters after their incomplete removal (and possible degradation) in the wastewater treatment plants (WWTP). A large amount of data about parent PhACs in water has been reported in the literature in the last decade; however, there is a lack of information about the presence of their metabolites and transformation/degradation products (TPs). Recent publications report that PhACs found in water are only the “top of the iceberg” in relation to the environmental impact associated to the consumption of PhACs. From a scientific-technological point of view, the overall study of PhACs is a challenge and requires to advance with respect to current knowledge and analytical capacities, considering several key aspects, such as the reliable identification and quantification of the compounds, the improvement of the removal efficiency by the WWTPs, the study of their behaviour in the environment (e.g. persistence and biodegradability), and the environmental risk assessment, considering not only the parent PhACs but also their transformation/metabolism products. In this work, it is intended to delve into this problem, presenting a detailed overview on metabolites and TPs of PhACs in environmental waters from the Mediterranean area. Analytical and environmental problems associated to the determination of these compounds are briefly commented, ending the paper with the main conclusions and expected future trends in relation to this field.     

氢氧根和烷氧基负离子含氧亲核试剂在富勒烯衍生化中的应用

富勒烯衍生化合物在有机光电、生物医学等领域表现出良好应用前景,相关的富勒烯衍生化方法研究引起了广泛关注,近年来发展迅速。富勒烯由于具有较强的缺电子性质,容易与亲核试剂反应。但相比于碳亲核试剂,含氧亲核试剂与富勒烯的反应发展缓慢,长期被忽略。我们结合本课题组最近开展的工作,对OH~-与MeO~-含氧亲核试剂参与的富勒烯衍生化反应的最新进展进行了总结,并对反应机理进行了较为详尽的阐述,为进一步扩展富勒烯衍生化方法提供参考。     

Determination of polybrominated diphenyl ethers in marine biological tissues using microwave-assisted extraction

Growing concern on the environmental impact of polybrominated diphenyl ethers (PBDEs) has created the need for rapid and quality assured analytical methods to quantify PBDEs in a spectrum of matrix types. This study presents the first validated method for the quantification of major PBDE congeners (47, 99 and 100) in marine biological tissues using microwave-assisted extraction (MAE). The recovery of polychlorinated biphenyls and various organochlorine pesticides has also been ascertained. Analytical accuracy, precision, limits of detection and cleanup efficiency were evaluated for PBDE congeners, and empirical data justifies the use of MAE for the extraction and analysis of PBDEs in biological matrices. MAE was also compared to Soxhlet extraction efficiency for PBDEs in the standard reference materials SRM2978 and SRM1588a and gave comparable results (<15% variation).     

An analytical method for determination of fullerenes and functionalized fullerenes in soils with high performance liquid chromatography and UV detection

Fullerenes are carbon-based nanomaterials expected to play a major role in emerging nanotechnology and produced at an increasing rate for industrial and household applications. In the last decade a number of novel compounds (i.e. fullerene derivatives) is being introduced into the market and specific analytical methods are needed for analytical purposes as well as environmental and safety issues. In the present work eight fullerenes (C60 and C70) and functionalized fullerenes (C60 and C70 exohedral-derivatives) were selected and a novel liquid chromatographic method was developed for their analysis with UV absorption as a method of detection. The resulting HPLC-UV method is the first one suitable for the analysis of all eight compounds. This method was applied for the analysis of fullerenes added to clayish, sandy and loess top-soils at concentrations of 20, 10 and 5 μg kg⁻¹ and extracted with a combination of sonication and shaking extraction. The analytical method limits of detection (LoD) and limits of quantification (LoQ) were in the range of 6–10 μg L⁻¹ and 15–24 μg L⁻¹ respectively for the analytical solutions. The extraction from soil was highly reproducible with recoveries ranging from 47 ± 5 to 71 ± 4% whereas LoD and LoQ for all soils tested were of 3 μg kg⁻¹ and 10 μg kg⁻¹ respectively. No significant difference in the extraction performance was observed depending of the different soil matrices and between the different concentrations. The developed method can be applied for the study of the fate and toxicity of fullerenes in complex matrices at relatively low concentrations and in principle it will be suitable for the analysis of other types of functionalized fullerenes that were not included in this work.     

富勒烯合成化学研究进展

富勒烯是一类由12个五元环和若干六元环组成的笼状分子, 自20世纪80年代中期被发现以来就以其独特的结构和新奇的性质而成为科学界研究的热点, 25年来, 无论在基础研究还是在实际应用领域都有了长足的进步, 人们在发展富勒烯合成新方法和寻找富勒烯新结构方面做了大量的工作。本文对富勒烯的各种宏量合成方法进行了回顾, 并概述了迄今已发表的60余种富勒烯新结构,包括各种富勒烯空笼、内嵌富勒烯、富勒烯笼外修饰衍生物及氮杂富勒烯等结构。     

Overview of analysis of carcinogenic and/or mutagenic metals in biological and environmental samples. I. Arsenic, beryllium, cadmium, chromium and selenium

One of the most dangerous and pernicious forms of pollution arises from the potential mobilization of a spectrum of toxic trace metals and metalloids in our environment. Among the most important elements in this regard are arsenic, beryllium, cadmium, chromium and selenium whose adverse toxic effects are now well recognized including their carcinogenicity and/or mutagenicity. These agents (and their derivatives) can be widely dispersed throughout the environment as a result of fossil fuel combustion, industrial and agricultural processes and natural processes. The trend for the immediate future appears to be of greater exposure to these metals not only as a result of generally increased usage patterns but also because of prospective enhanced use of fossil fuels for space heating and electricity generation. In order to more readily evaluate trends of human exposure as well as the toxicity, bioavailability, bioaccumulation and transport of these elements, sensitive analytical procedures are required for the determination of their various oxidation states (as well as their organic derivatives) in complex matrices such as those found in both environmental and biological samples. Hence, the principal objective of this overview is to highlight the more recent trends and state-of-the-art methodologies for the determination of arsenic, beryllium, cadmium, chromium and selenium (in their various forms) in environmental compartments such as air, water, soil and in human tissues (primarily blood, urine, and milk). Techniques to be discussed primarily include atomic absorption spectrometry, neutron activation analysis, gas chromatography, differential pulse polarography and electrochemical analysis. The importance of quality control and differentiation according to speciation will also be stressed.     

Implications of the solvent effect in quantitative capillary gas chromatography of minor constituents in mixtures—a study using deuterium labeled and unlabeled benzofluoranthenes

Perdeuterated benzofluoranthenes have slightly shorter retention times than their equivalent unlabeled forms in a DB-5 capillary column. Under column overload conditions, perdeuterated benzofluoranthenes in moderate multifold excesses, which might be encountered in their use as internal standards and carriers for quantitative analysis, are seen to exhibit both normal and reverse solvent effects on their close eluting congeners. In some cases the effects may be used to advantage by knowledgeable analysts, but for the ignorant and unwary the effects can lead to serious errors in identification and quantification.     

Extraction and high-performance liquid chromatographic analysis of C60, C70, and [6,6]-phenyl C61-butyric acid methyl ester in synthetic and natural waters

Studies have shown that C(60) fullerene can form stable colloidal suspensions in water that result in C(60) aqueous concentrations many orders of magnitude above C(60)'s aqueous solubility; however, quantitative methods for the analysis of C(60) and other fullerenes in environmental media are scarce. Using a 80/20v/v toluene-acetonitrile mobile phase and a 4.6mmx150mm Cosmosil 5mu PYE column, C(60), C(70), and PCBM ([6,6]-phenyl C(61)-butyric acid methyl ester) were fully resolved. Selectivity factors (alpha) for C(60) relative to PCBM and C(70) relative to C(60) were 3.18 and 2.19, respectively. The best analytical wavelengths for the fullerenes were determined to be 330, 333, and 333nm with log molar absorption coefficients (logvarepsilon) of 4.63, 4.82, and 4.60 for PCBM, C(60), C(70), respectively. Extraction and quantitation of all three fullerenes in aqueous suspensions over a range of pH (4-10) and ionic strengths were very good. Whole-method quantification limits for ground and surface suspensions were 2.87, 2.48, and 6.54mug/L for PCBM, C(60), and C(70), respectively.     

A biophysical perspective of understanding nanoparticles at large

In this article we present a biophysical perspective that describes the fate of nanoparticles in both the aqueous phase and in living systems. Specifically, we show the correlations between the physicochemistry of fullerenes and their uptake, translocation, transformation, transport, and biodistribution in mammalian and plant systems, at the molecular, cellular, and whole organism level. In addition to fullerenes and their structural derivatives, we describe the biological and environmental implications and applications of the condensed matter of carbon nanotubes and quantum dots, and the soft condensed matter of plastic and dendrimers. The main purpose of this article is to demonstrate the vast opportunities and unique advantages of applying experimental and simulation biophysics to the nascent research field of understanding nanoparticles at large.     

High resolution gas chromatography of technical chlordane with electron capture- and mass selective detection

Summary High resolution gas chromatography with electron capture- and mass selective detection was used for the characterization of the constituents of the technical cyclodiene pesticide chlordane. Degradation reactions were used for confirmation of tentatively identified chlordane components in environmental samples. The pattern of chlordane congeners in cod liver from Norway is described. Compared to the technical product the nonachloro congeners are unchanged in their relative occurrence. Among the octachloro congeners, trans-chlordane is diminished relative to the cis-chlordane.

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Hochauflösende Gas-Chromatographie von technischem Chlordan mit Hilfe der Elektroneneinfang- und massenselektiven Detektion

     

同位素稀释-气相色谱-串联质谱法测定土壤中的指示性毒杀芬

建立了土壤样品中指示性毒杀芬Parlar No.26 (P26)、Parlar No.50 (P50)和Parlar No.62 (P62)的同位素稀释-气相色谱-串联质谱(ID-GC-MS/MS)的分析方法。土壤样品使用压力溶剂萃取装置(PLE)提取,以丙酮-正己烷(1:1, v/v)混合溶液为提取溶剂;提取液依次经由多层酸性硅胶柱和活化硅胶柱净化;洗脱液经氮吹浓缩至20 μL后,利用GC-MS/MS的多反应监测(MRM)模式进行定性和定量。结果表明本方法可对样品中的P26、P50和P62进行分析,相对标准偏差(RSD)小于11%,回收率可以达到55%～110%;P26、P50和P62的仪器检出限分别为3.0、3.0和6.0 pg。将该方法用于某地区农田表层土壤中3种指示性毒杀芬的检测,其中P26的含量为0.17 ng/g、P50为0.08 ng/g、P62为0.09 ng/g。此方法适用于土壤样品中指示性毒杀芬的分析。     

Tailoring enzyme-rich environmental DNA clones: a source of enzymes for generating libraries of unnatural natural products

A detailed bioinformatics analysis of six glycopeptide biosynthetic gene clusters isolated from soil environmental DNA (eDNA) megalibraries indicates that a subset of these gene clusters contains collections of tailoring enzymes that are predicted to result in the production of new glycopeptide congeners. In particular, sulfotransferases appear in eDNA-derived gene clusters at a much higher frequency than would be predicted from the characterization of glycopeptides from cultured Actinomycetes . Enzymes found on tailoring-enzyme-rich eDNA clones associated with these six gene clusters were used to produce a series of new sulfated glycopeptide derivatives in both in vitro and in vivo derivatization studies. The derivatization of known natural products with eDNA-derived tailoring enzymes is likely to be a broadly applicable strategy for generating libraries of new natural product variants.     

富勒烯和富勒烯衍生物中的Stone-Wales旋转

Stone-Wales旋转是富勒烯异构化的基本方式,了解其特征和规律对于理解富勒烯和富勒烯衍生物的形成至关重要.本文采用密度泛函理论方法系统研究了富勒烯和富勒烯衍生物的Stone-Wales旋转.结果显示,富勒烯异构体趋向于从高B55键(两个五元环共用的边)结构向低B55结构转化,满足独立五元环原则的结构或具有低B55键数的异构体在热力学上更为有利.相反,对于富勒烯衍生物,具有更多B55的异构体不仅在热力学上更有利,而且从动力学角度讲,从满足独立五元环原则的结构向不满足的结构的转变比相反过程更容易.这些结果可以解释目前的相关实验事实,暗示了富勒烯衍生物可能是先衍生化后异构化而形成.     

DNA aptamers for selective identification and separation of flame retardant chemicals

Polybrominated diphenyl ethers (PBDEs) are group of chemicals which are representative persistent organic pollutants (POPs) and used as brominated flame retardants for many consumer products. PBDEs were phased out since 2009 but are still frequently observed in various environmental matrices and human body. Here, we report ssDNA aptamers which bind to BDE47, one of the PBDE congeners commonly found in various environmental matrices, and show affinity to other major tri-to hepta- BDE congeners. The PBDE specific aptamers were isolated from random library of ssDNA using Mag-SELEX. Two out of 15 sequences, based on their alignment and hairpin loop structures, were chosen to determine dissociation constant with BDE47 and showed from picomolar to nanomolar affinities (200 pM and 1.53 nM). The aptamers displayed high selectivity to the original target, BDE47, and implying general specificity to PBDE backbone with varying affinities to other congeners. Further, we showed that the use of two aptamers together could enhance the separation efficiency of BDE47 and other BDE congeners when dissolved in a solvent compared to use of single aptamer. These aptamers are expected to provide a tool for preliminary screening or quick separation of PBDEs in environmental samples prior to trace quantitative analysis.     

Aminoglycoside antibiotics may interfere with microbial amino sugar analysis

The amino sugars (e.g., glucosamine, galactosamine, mannosamine, muramic acid) in soils are frequently employed as biomarkers of microbial residues. The analysis of amino sugars in environmental matrices, however, is expected to be more complicated than their determination in isolated microbial cells. In this study, we employed a widely used protocol for amino sugar analysis, and found that some aminoglycoside antibiotics interfere with amino sugar quantification in vitro. The method converts the aminoglycosides to compounds that coelute with the aldononitrile acetate derivatives of the amino sugars. Specifically, streptomycin significantly interferes with muramic acid analysis, and kanamycin, tobramycin and amikacin hamper glucosamine measurement. Mass spectrometry confirmed that the interfering compounds from aminoglycosides are not actually genuine microbial amino sugar monomers (bacterial muramic acid or fungal glucosamine), and are most likely to be N-methyl glucosamine or 3-amino-3-deoxy-glucopyranose. In contrast to their effects on muramic acid and glucosamine analyses, aminoglycosides do not interfere with galactosamine and mannosamine quantification. The few data that exist on the environmental occurrence of aminoglycoside antibiotics suggest they occur at only trace levels. Our findings may have implications for the qualitative and quantitative validity of results from amino sugar assays in some context. Application of the aldononitrile acetate derivatization method to samples (especially in selective microbial cultures using aminoglycosides as inhibitors) requires that potential interference be evaluated.