基礎講座 電気泳動(Electrophoresis)

Dissociation constants of organic acids in aqueous solution : G. Kortüm, W. Vogel and K. Andrussow, Butterworths, London, 1961, xxii + 347 pages, £ 2.108

近日，原热电科学仪器公司和飞世尔科学公司在北京联合召开新闻发布会，宣布双方的合并完成，新的实体Thermo Fisher Scientific正式宣告成立。此次新闻发布会是Thermo Fisher成立后的全球第一个新闻发布会，同时也揭开了公司总裁兼首席执行官Marijn E Dekkers中国之行的序幕。当天，有近60家来自仪器、医疗、生物、化工、环境、医药和电子等行业媒体及经济类媒体出席了发布会。

科学仪器公司Thermo Fisher Scientific正式起航

Glyphosate [N-(phosphonomethyl)glycine] is the most frequently used herbicide to date. Due to its indiscriminate use, it has become a globally occurring pollutant of surface waters. A biosensor for glyphosate is described here that consists of a carbon nano-onion/tyrosinase conjugate immobilized in a chitosan matrix on a screen-printed electrode. The analytical principle is based on the inhibition of the enzyme tyrosinase by glyphosate. L-DOPA is used as the enzyme substrate. The presence of the carbon nano-onions has a beneficial effect on the sensitivity of the assay. Glyphosate can be amperometrically quantified in the 0.015 to 10 μM concentration range and with a 6.5 nM (1.1 μg L−1) detection limit. The biosensor is stable more than 2 months at 4 °C. It was applied to the detection of glyphosate in water and soil samples taken from irrigation of a rice field after aerial application. Results were in good agreement with data obtained by a commercial ELISA.

Amperometric biosensor for glyphosate based on the inhibition of tyrosinase conjugated to carbon nano-onions in a chitosan matrix on a screen-printed electrode.

Glyphosate (N-(phosphonomethyl)glycine) is the most frequently used broad-spectrum herbicide worldwide. Its mechanism of action is based on the inhibition of an enzyme that is essential to plant growth. Its intensive use has caused global contamination to occur, which has not only affected the ecosystems, but even food and other objects of common use. Thus, there is a pronounced need for developing analytical methods for glyphosate determination in different matrices. Here, an electrochemical competitive immunoassay, based on the use of antibody-modified magnetic particles, has been developed. Tetramethylbenzidine (TMB) has been used as an enzymatic substrate. The extent of the affinity reaction has been achieved by monitoring the current value, due to the reduction of the enzymatic product. A disposable screen-printed electrochemical cell has been used. The calibration curve has been recorded in the 0–10,000 ng/L concentration range, with a detection limit of 5 ng/L and quantification limit of 30 ng/L. The electrochemical immunoassay has also been applied to the analysis of spiked beer samples.

Glyphosate Determination by Coupling an Immuno-Magnetic Assay with Electrochemical Sensors

Since its introduction in 1974, the herbicide glyphosate has experienced a tremendous increase in use, with about one million tons used annually today. This review focuses on sensors and electromigration separation techniques as alternatives to chromatographic methods for the analysis of glyphosate and its metabolite aminomethyl phosphonic acid. Even with the large number of studies published, glyphosate analysis remains challenging. With its polar and depending on pH even ionic functional groups lacking a chromophore, it is difficult to analyze with chromatographic techniques. Its analysis is mostly achieved after derivatization. Its purification from food and environmental samples inevitably results incoextraction of ionic matrix components, with a further impact on analysis derivatization. Its purification from food and environmental samples inevitably results in coextraction of ionic matrix components, with a further impact on analysis and also derivatization reactions. Its ability to form chelates with metal cations is another obstacle for precise quantification. Lastly, the low limits of detection required by legislation have to be met. These challenges preclude glyphosate from being analyzed together with many other pesticides in common multiresidue (chromatographic) methods. For better monitoring of glyphosate in environmental and food samples, further fast and robust methods are required. In this review, analytical methods are summarized and discussed from the perspective of biosensors and various formats of electromigration separation techniques, including modes such as capillary electrophoresis and micellar electrokinetic chromatography, combined with various detection techniques. These methods are critically discussed with regard to matrix tolerance, limits of detection reached, and selectivity.

Glyphosate analysis using sensors and electromigration separation techniques as alternatives to gas or liquid chromatography.

In this study, we developed and validated a CE-TOF-MS method for the quantification of glyphosate (N-(phosphonomethyl)glycine) and its major degradation product aminomethylphosphonic acid (AMPA) in different samples including beer, media from toxicological analysis with Daphnia magna, and sorption experiments. Using a background electrolyte (BGE) of very low pH, where glyphosate is still negatively charged but many matrix components become neutral or protonated, a very high separation selectivity was reached. The presence of inorganic salts in the sample was advantageous with regard to preconcentration via transient isotachophoresis. The advantages of our new method are the following: no derivatization is needed, high separation selectivity and thus matrix tolerance, speed of analysis, limits of detection suitable for many applications in food and environmental science, negligible disturbance by metal chelation. LODs for glyphosate were < 5 μg/L for both aqueous and beer samples, the linear range in aqueous samples was 5–3000 μg/L, for beer samples 10–3000 μg/L. For AMPA, LODs were 3.3 and 30.6 μg/L, and the linear range 10–3000 μg/L and 50–3000 μg/L, for aqueous and beer samples, respectively. Recoveries in beer samples for glyphosate were 94.3–110.7% and for AMPA 80.2–100.4%. We analyzed 12 German and 2 Danish beer samples. Quantification of glyphosate and AMPA was possible using isotopically labeled standards without enrichment, purification, or dilution, only degassing and filtration were required for sample preparation. Finally, we demonstrate the applicability of the method for other strong acids, relevant in food and environmental sciences such as N-acetyl glyphosate, N-acetyl AMPA (present in some glyphosate resistant crop), trifluoroacetic acid, 2-methyl-4-chlorophenoxyacetic acid, glufosinate and its degradation product 3-(methylphosphinico)propionic acid, oxamic acid, and others.

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Capillary electrophoresis-mass spectrometry for the direct analysis of glyphosate: method development and application to beer beverages and environmental studies.

Electrophoretic separations are of growing interest to tackle complex analytical challenges. Nevertheless, capillary electrophoresis, as the most common mode, still suffers from insufficient detection limits due to low capillary loadability. ITP is of growing interest as preconcentration method for capillary electrophoresis and is also interesting to be applied as an independent analytical method. While mass spectrometric detection is common for capillary electrophoresis, the combination of ITP with MS is still a niche technique. In this work, we want to give an overview on isotachophoretic effects in CE-MS and ITP-MS methods, as well as coupling techniques of ITP with CE-MS. The challenges and possibilities associated with mass spectrometric detection in ITP and its coupling to capillary electrophoresis are critically discussed.

Challenges and applications of isotachophoresis coupled to mass spectrometry: A review

13C assimilation as well as functional gene abundance and expression elucidate the biodegradation of glyphosate in a field experiment.

BACKGROUND
Analytical constraints complicate environmental monitoring campaigns of the herbicide glyphosate and its major degradation product aminomethylphosphonic acid (AMPA): their strong sorption to soil minerals require harsh extraction conditions. Coextracted matrix compounds impair downstream analysis and must be removed before analysis.


RESULTS
A new extraction method combined with subsequent capillary electrophoresis-mass spectrometry for derivatization-free analysis of glyphosate and AMPA in soil and sediment was developed and applied to a suite of environmental samples. It was compared to three extraction methods from literature. We show that no extraction medium reached 100% recovery. The new phosphate-supported alkaline extraction method revealed 1) high recoveries of 70-90% for soils and aquatic sediments, 2) limits of detections below 20 μg kg-1 , and 3) a high robustness, because impairing matrix components (trivalent cations and humic acids) were precipitated prior to the analysis. Soil and sediment samples collected around Tübingen, Germany, revealed maximum glyphosate and AMPA residues of 80 and 2100 μg kg-1 , respectively, with residues observed along a core of lake sediments. Glyphosate and/or AMPA were found in 40% of arable soils and 57% of aquatic sediment samples.


CONCLUSION
In this work, we discuss soil parameters that influence (de)sorption and thus extraction. From our results we conclude that residues of glyphosate in environmental samples are easily underestimated. With its possible high throughput, the method presented here can resolve current limitations in monitoring campaigns of glyphosate by addressing soil and aquatic sediment samples with critical sorption characteristics. This article is protected by copyright. All rights reserved.

Benedikt Wimmer

Papers

Glyphosate analysis using sensors and electromigration separation techniques as alternatives to gas or liquid chromatography.

Capillary electrophoresis-mass spectrometry for the direct analysis of glyphosate: method development and application to beer beverages and environmental studies.

Challenges and applications of isotachophoresis coupled to mass spectrometry: A review

13C assimilation as well as functional gene abundance and expression elucidate the biodegradation of glyphosate in a field experiment.

Phosphate addition enhances alkaline extraction of glyphosate from highly sorptive soils and aquatic sediments.