WO2020186995A1 - 一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法 - Google Patents

一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法 Download PDF

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WO2020186995A1
WO2020186995A1 PCT/CN2020/077103 CN2020077103W WO2020186995A1 WO 2020186995 A1 WO2020186995 A1 WO 2020186995A1 CN 2020077103 W CN2020077103 W CN 2020077103W WO 2020186995 A1 WO2020186995 A1 WO 2020186995A1
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hydrogen sulfide
smart device
sample
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唐盛
祁桐
沈薇
许孟婵
许孟媛
朱安妮
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江苏科技大学
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
    • G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
    • G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/02—Food
    • G01N33/04—Dairy products
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N1/22—Devices for withdrawing samples in the gaseous state
    • G01N1/2202—Devices for withdrawing samples in the gaseous state involving separation of sample components during sampling
    • G01N1/2214—Devices for withdrawing samples in the gaseous state involving separation of sample components during sampling by sorption
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
    • G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
    • G01N21/783—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour for analysing gases
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N1/22—Devices for withdrawing samples in the gaseous state
    • G01N1/2202—Devices for withdrawing samples in the gaseous state involving separation of sample components during sampling
    • G01N1/2214—Devices for withdrawing samples in the gaseous state involving separation of sample components during sampling by sorption
    • G01N2001/2217—Devices for withdrawing samples in the gaseous state involving separation of sample components during sampling by sorption using a liquid
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N1/22—Devices for withdrawing samples in the gaseous state
    • G01N1/2226—Sampling from a closed space, e.g. food package, head space
    • G01N2001/2229—Headspace sampling, i.e. vapour over liquid

Definitions

  • the invention relates to a method for measuring hydrogen sulfide by using a headspace single drop liquid phase microextraction method and intelligent equipment colorimetric determination, and belongs to the technical field of hydrogen sulfide detection.
  • Hydrogen sulfide is a colorless acid gas with a unique smell of rotten eggs. It is understood that H 2 S can be produced in the food processing industry, coal gasification plants, and crude oil refining processes. It is the third toxic gas signal molecule after nitric oxide and carbon monoxide. It has important pathophysiological effects in many cardiovascular diseases, such as cardiac ischemia, vasodilation and neuromodulation. More and more evidence shows that abnormal levels of H 2 S are closely related to diabetes, Alzheimer's disease and Down syndrome. When the concentration of hydrogen sulfide rises to 15ppm, people exposed to it will lose consciousness, accompanied by apnea and inactivation of the olfactory system. Concentrations exceeding 320ppm can cause death. The spoilage process of food such as eggs and milk also produces H 2 S. Therefore, rapid and sensitive detection of H 2 S is of great significance for early warning of food spoilage or contamination events.
  • nanomaterials to detect H 2 S in gas phase and biological systems such as carbon nanostructures, metal nanoparticles, metal oxide nanoparticles and quantum dots.
  • These materials have high selectivity as miniature sensors, can be applied in real time, and other outstanding advantages.
  • silver/gold nanomaterials have been reported to suppress and shift surface plasmon resonance signals, or as electrochemical sensors to detect H 2 S in serum or HeLa cells.
  • the nanoscale sensor is directly mixed with the sample, and there is serious matrix interference.
  • the potential matrix effect of biological samples can lead to low selectivity to H 2 S. Therefore, it is necessary to adopt new analysis methods to eliminate matrix effects.
  • Single drop microextraction is a liquid phase microextraction method, which is an effective liquid phase and gas analysis analyte extraction method.
  • Headspace (HS)-SDME is most effective against volatile compounds. Because volatile compounds can be separated from the sample into the headspace, there is no contact between the solvent and the sample during the extraction process. Therefore, the interference problem of complex systems is completely avoided.
  • analyzing individual droplets is a challenge. In traditional HS-SDME, especially when using microsyringes for analysis, it is generally necessary to combine gas chromatography or liquid chromatography to analyze droplets. Otherwise, it is difficult to use other analysis methods.
  • many instrument companies have developed several nanometer droplet detectors for analyzing extractant droplets. However, these detectors are application-specific, relatively expensive, and inconvenient to obtain.
  • the present invention provides a method for colorimetric determination of hydrogen sulfide by using a headspace single drop liquid phase microextraction method and smart equipment.
  • a method for the colorimetric determination of hydrogen sulfide by using a single droplet liquid phase microextraction method in the headspace and a smart device including the following steps:
  • the silver-gold core-shell triangular nanosheet (Ag@Au TNS) is used as the nano detection probe, combined with the analysis method of headspace single drop microextraction (HS-SDME), so that the H 2 S volatilized in the sample to be tested is nanometer
  • HS-SDME headspace single drop microextraction
  • the detection probe is specifically extracted, and the extracted sample is taken, and H 2 S is detected with the help of smart device camera function and color picking software.
  • the preparation method of the silver-gold core-shell triangular nanosheet includes the following steps:
  • step (2) the PVP, ascorbic acid, silver nitrate solution, and sodium citrate solution can be used in the usual amounts.
  • the volume ratio of PVP, diethylamine and ascorbic acid in step (3) is (1-9):(0.25-1.25):1, and the components of the gold-containing solution include PVP, KI, HAuCl 4 and ultrapure
  • the ratio of the total mass of water, Ag and Au is (10-4):1.
  • the method for specific extraction of the nano detection probe includes the following steps:
  • the method for detecting H 2 S with the help of smart device camera function and color picking software includes the following steps:
  • RGB value is directly provided by the image taken by the color picking software
  • LOD refers to the lowest detection limit
  • ⁇ black is the standard deviation of the blank solution
  • k is the slope of the standard curve.
  • the smart devices include smart phones and tablet computers.
  • the color picking software is EKColorPicker software, color picker software or ChemEye.
  • smart devices such as smart phones
  • the use of smart devices provides an interesting platform for diagnosis and environmental monitoring.
  • most of the analysis methods developed on the smart device are colorimetry and macroscopic feature imaging.
  • the color can be analyzed and the relationship corresponding to the analyzed concentration can be found.
  • the smart device camera is also suitable for taking images of a single solvent droplet, just like what exists in SDME.
  • the method of the present invention utilizes the extraction agent Ag@Au TNS pre-added to SDME to be inhibited by the ultraviolet-visible (UV-vis) signal caused by H 2 S etching, and measures the H 2 S concentration.
  • the coating of the gold layer not only ensures the high stability of the nanomaterials, but also improves the selectivity to H 2 S.
  • the HS-SDME method has a simple process and only a drop of solvent is required to complete the analysis.
  • the present invention adopts the smart device colorimetry method, the detection limit is about 65nM, and the linear range is 0.1 ⁇ M-100 ⁇ M.
  • the established method can be applied to the determination of actual samples such as egg white and milk in opaque samples
  • the H2S has the advantages of less flow, simple operation and high detection efficiency.
  • Figure 1 Mechanism diagram of H 2 S detection method using silver-gold core-shell triangular nanosheets headspace single drop liquid phase microextraction
  • Figure 2 TEM images of prepared Ag TNS and Ag@Au TNS with three different gold layer thicknesses: I (B and C), II (D, E and F), III (G and H); where A is Ag TEM image of TNS; B and C are TEM images of Ag@Au TNS with I layer gold; D, E and F are TEM images of Ag@Au TNS with II layer gold; G and H are Ag@Au with III layer gold TEM image of TNS.
  • Figure 3 TEM images of Ag@Au TNS incubated in 100 ⁇ M H 2 S for 5, 10, 15, 20 min and EDX mapping before and after the reaction. Among them, 1) is the TEM image of Ag@Au TNS incubated in 100 ⁇ M Na 2 S for 5, 10, 15, and 20 minutes; 2) is the EDX mapping of Ag@Au TNS before 100 ⁇ M H 2 S reaction; 3) is Ag@ EDX mapping of Au TNS after 100 ⁇ M H 2 S reaction.
  • Figure 4 The UV-Vis absorption spectrum of Ag@Au TNS after being exposed to 100 ⁇ M Na 2 S HS-SDME for 20 minutes.
  • Figure 5 Ag@Au TNS in 100 ⁇ M Na 2 S, the incubation temperature is from 15 °C to 35 °C; the pH of the target solution is from 5 to 9; the incubation time is optimized from 0 min to 20 min.
  • FIG. 7 Ag TNS, Ag@Au TNS I, II, III, respectively, 100 ⁇ M Na 2 S solution (blue) and 100 ⁇ M Na 2 SO 3 solution (red) after 20 minutes of HS-SDME absorption spectrum changes.
  • Figure 8 Calibration curve of H 2 S UV detection. Conditions: 100 ⁇ M H 2 S, pH 6 , 30°C, incubate for 20 min;
  • Figure 9 Calibration curve of H 2 S smart phone nano colorimetric detection. Conditions: 100 ⁇ M H 2 S, pH 6 , 30°C, incubate for 20 min;
  • Figure 11 The concentration curve of hydrogen sulfide when milk is stored at 4°C and 25°C in 10 days;
  • Figure 12 The concentration curve of hydrogen sulfide when eggs are stored at 4°C and 25°C in 10 days.
  • Example 1 Method for colorimetric determination of hydrogen sulfide using headspace single drop liquid phase microextraction method and smart equipment
  • Figure 2 shows the TEM images of the prepared Ag TNS and Ag@Au TNS with three different gold layer thicknesses. It can be observed from Figure 2A that the average side length of Ag TNS is about 55 nm. The thickness of the three different Au layers of Ag@Au TNS are 0.8nm, 1.58nm and 2.7nm, respectively, as shown in Figure 2B-H.
  • FIG. 1 it is a mechanism diagram of a method for detecting H 2 S after single-drop liquid phase micro-extraction using silver-gold core-shell triangular nanosheets.
  • the droplets With the help of the surface tension of water and intermolecular forces, the droplets will not easily fall from the top unless the centrifuge tube vibrates violently. Therefore, it is feasible to perform SDME in this way.
  • the cap on the centrifuge tube quickly and gently, and let the centrifuge tube stand for 20 minutes to ensure that the H 2 S is released from the sample and is fully extracted by the extract droplets.
  • SDME carefully open the lid of the centrifuge tube and transfer 3.0 ⁇ L of extractant to the NanoPhotometer UV-Vis spectrophotometer using a pipette to analyze its UV-visible light signal.
  • SNC the photo of the droplet is taken by the smartphone camera.
  • the RGB (red, green, blue) colors of the image are analyzed by EKColorPicker.
  • the R (red) value of the image is directly provided by the EKColorPicker software to indicate the intensity of the color.
  • the added PVP, ascorbic acid and sodium citrate need to be added at intervals of 10 minutes.
  • the concentration of silver nitrate in the mixed solution of silver nitrate and sodium citrate is 0.75 mM, and the concentration of sodium citrate is 1.13 mM.
  • the gold-containing solution is: 400 ⁇ L PVP (vinylpyrrole monomer concentration is 0.5M), 80 ⁇ L KI (0.2M), 20 ⁇ L HAuCl 4 (0.25M), 3 mL ultrapure water. And add at a rate of 0.05mL/min.
  • the NaBH 4 (0.1M): It is prepared from ice water and is prepared for immediate use.
  • Measured data The RGB value is directly provided by the image taken by the color picking software, and the standard curve is drawn by measuring the data of the R (red) value.
  • LOD refers to the lowest detection limit
  • ⁇ black is the standard deviation of unreacted Ag@Au TNS
  • k is the slope of the standard curve.
  • Figure 5 is an optimized experiment diagram. The optimal experimental conditions are pH 6, 30°C, and extraction for 20 minutes.
  • Figures 8 and 9 are the calibration curve of H 2 S UV detection and the calibration curve of H 2 S smart phone nano colorimetric detection. The detection limits are about 7nM and 65nM, respectively, and the linear ranges are 10nM-10 ⁇ M and 0.1 ⁇ M-100 ⁇ M.
  • Figure 10 shows the UV spectra corresponding to different concentrations of H 2 S.
  • the milk sample of fresh H 2 S were measured and continuously monitored for 10 days.
  • the fresh milk purchased directly from the supermarket is placed in two beakers for storage, and stored in a refrigerator at 4°C and at room temperature 25°C.
  • An ultraviolet-visible spectrophotometer was used to measure the experimental data, and the trend chart of the H 2 S in the milk within 10 days was obtained, as shown in Figure 11.
  • these samples were added with 0.02 ⁇ M, 0.2 ⁇ M, and 2 ⁇ M H 2 S and detected by UV-Vis spectrophotometer; respectively added with 0.2 ⁇ M, 2 ⁇ M and 20 ⁇ M, and H 2 S was detected by smart phone nanocolorimetry.
  • the egg sample H 2 S were measured and continuously monitored for 10 days. Eggs purchased directly from the supermarket were stored in a refrigerator at 4°C and at room temperature 25°C. Open a skylight with a diameter of 5mm at one end of the egg shell. An ultraviolet-visible spectrophotometer was used to measure the experimental data, and a trend chart of the H 2 S in the egg within 10 days was obtained, as shown in Figure 12. In order to evaluate the matrix effect, these samples were added with 0.02 ⁇ M, 0.2 ⁇ M, and 2 ⁇ M H 2 S and detected by UV-Vis spectrophotometer; respectively added with 0.2 ⁇ M, 2 ⁇ M and 20 ⁇ M, and H 2 S was detected by smart phone nanocolorimetry. The results are shown in Tables 3 and 4, respectively, and a good recovery rate (95-104%) was obtained.

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Abstract

一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢(H 2S)的方法，包括以下步骤：以银-金核壳三角形纳米薄片(Ag@Au TNS)作为纳米检测探针，结合顶空单滴微萃取(HS-SDME)的分析方法，使待测样品中挥发出的H 2S被纳米检测探针特异性萃取，取萃取后的样本，借助智能设备拍照功能和取色软件对H 2S进行检测。相对于现有技术，采用智能设备比色法，检测极限为65nM左右，线性范围为0.1μM-100μM，所建立的方法能够应用于测定实际样品如蛋清及牛奶等不透明样本中的H 2S，具有流程少、操作简单、检测效率高等优点。

Description

一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法 技术领域
本发明涉及一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法，属于硫化氢检测技术领域。
背景技术
硫化氢(H 2S)是一种无色酸性气体，具有臭鸡蛋特有的气味。据了解，食品加工业，煤气化厂，以及原油炼制过程中均可产生H 2S。它是继一氧化氮和一氧化碳之后的第三种有毒气体信号分子。在许多心血管疾病中具有重要的病理生理学作用,如心脏缺血，血管舒张和神经调节等。越来越多的证据表明，H 2S的异常水平与糖尿病、阿尔茨海默病和唐氏综合症密切相关。当硫化氢浓度上升到15ppm时，暴露于其中的人将失去意识，并伴有呼吸暂停和嗅觉***失活，浓度超过320ppm可导致死亡。鸡蛋、牛奶等食物的变质过程也会产生H 2S。因此，快速、灵敏地检测H 2S对于预警食品变质或污染事件具有重要意义。
迄今为止，报道了许多经典测定H 2S的方法，如气相色谱(GC)、电化学、化学发光等。一般来说，上述方法都是测定H 2S较好的自动化仪器方法。然而，它们需要中等复杂的仪器和繁琐的预处理过程，以及气体(氦或氮)作为流动相和参考气体混合物进行分析***校准。荧光法也被广泛应用于生物样品中H 2S的检测和实时成像。近年来，荧光检测方法被广泛应用于H 2S检测和实时成像。但该技术在实际应用中仍存在局限性，如低选择性、低光稳定性和难以分离未反应物质等。
近年来，越来越多的研究集中在利用纳米材料在气相和生物***中检测H 2S，如碳纳米结构、金属纳米颗粒、金属氧化物纳米颗粒和量子点。这些材料作为微型传感器具有高选择性，可以实时应用，以及其他突出优点。例如，银/金纳米材料已经被报道通过表面等离子体共振信号的抑制和移位，或作为电化学传感器检测血清或HeLa细胞中的H 2S。然而，在这些方法中，纳米级传感器直接与样品混合，存在严重的基质干扰。生物样品的潜在基质效应会导致对H 2S的选择性较低。因此，有必要采用新的分析方法来消除基体效应。
单滴微萃取(SDME)是一种液相微萃取方法，它是一种有效的液相和气体分析的分析物萃取方法。顶空(HS)-SDME对挥发性化合物最为有效。因为挥发性化合物可以从样品中分离到顶空，萃取过程中溶剂和样品之间没有任何接触。因此，完全避免了复杂 ***的干扰问题。然而，分析单个液滴是一个挑战。在传统的HS-SDME中，尤其是使用微注射器进行分析时，一般要结合气相色谱法或液相色谱法对液滴进行分析。否则，很难使用其他分析方法。近年来，许多仪器公司研制了几种纳米液滴探测器，用于分析萃取剂液滴。然而，这些检测器是特殊应用的，相对昂贵，并且获取也并不方便。
发明内容
发明目的：为了解决上述技术问题，本发明提供了一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法。
技术方案：为了实现上述目的，本发明采用以下技术方案：
一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法，包括以下步骤：
以银-金核壳三角形纳米薄片(Ag@Au TNS)作为纳米检测探针，结合顶空单滴微萃取(HS-SDME)的分析方法，使待测样品中挥发出的H 2S被纳米检测探针特异性萃取，取萃取后的样本，借助智能设备拍照功能和取色软件对H 2S进行检测。
作为优选：
所述银-金核壳三角形纳米薄片的制备方法包括以下步骤：
(1)取硝酸银，柠檬酸钠和双氧水加入到去离子水中搅拌，之后快速加入NaBH 4，同时停止搅拌，一定时间后离心，清洗，即得银三角形纳米薄片(Ag TNS)，备用；
(2)将上述Ag TNS重新分散于去离子水中，加入PVP和抗坏血酸，随后依次滴加硝酸银溶液、柠檬酸钠溶液以及硝酸银和柠檬酸钠的混合溶液，混合均匀后，经离心后用去离子水清洗。
(3)通过再次加入PVP、二乙胺、抗坏血酸以及含金溶液，使金层沉淀在Ag TNS表面，最后将产物离心，去离子水洗涤，去除AgNO 3沉淀和多余的PVP，即得所述银-金核壳三角形纳米薄片(Ag@Au TNS)。
进一步优选，步骤(1)中硝酸银，柠檬酸钠和双氧水的摩尔比为1:(10-20):(200-360)，混合溶液与NaBH 4的体积比＝(1.3-2.5):1。
进一步优选，步骤(2)中PVP和抗坏血酸、硝酸银溶液、柠檬酸钠溶液按照常规的用量即可。
进一步优选，步骤(3)中所述PVP、二乙胺和抗坏血酸的体积比为(1-9):(0.25-1.25):1，含金溶液的成分包括PVP、KI、HAuCl 4和超纯水，Ag和Au的总物 质量之比为(10-4)：1。
所述纳米检测探针特异性萃取的方法包括以下步骤：
将待测样品加入带盖容器中，取下盖子，将微量Ag@Au TNS溶液滴到盖子内表面，随后将盖子盖在到容器上，静置一段时间，确保H 2S从样品中释放出来后被萃取液滴充分提取，结束后，打开盖子，其盖内表面液滴即为所述萃取后的样本。
所述借助智能设备拍照功能和取色软件对H 2S进行检测的方法包括以下步骤：
测取数据：RGB的值由取色软件通过拍摄的图像直接提供
绘制标曲：将计算得的RGB值的差值作为纵坐标，浓度的对数为横坐标得出标准曲线。
读取数据：通过得到的标准曲线，根据LOD＝3δblack/k，得出最低检测限浓度。其中LOD是指最低检测限，δblack是空白溶液的标准偏差，k是标准曲线的斜率。
所述智能设备包括智能手机、平板电脑。
所述取色软件为EKColorPicker软件、取色器软件或ChemEye。
本发明中所述采用智能设备(如智能手机)作为一种新型的分析设备，为诊断和环境监测提供了一个有趣的平台。基于智能设备摄像头这一优秀的彩色成像传感器，在智能设备上开发的分析方法大多是比色法和宏观特征成像。通过使用一些取色器软件，可以对颜色进行分析，找到与分析浓度相对应的关系。此外，该智能设备摄像头还适用于拍摄单个溶剂液滴的图像，就像SDME中存在的那样。
本发明方法利用预先加入SDME的萃取剂Ag@Au TNS被H 2S蚀刻引起的紫外可见(UV-vis)信号抑制，测量H 2S浓度。金层的包覆不仅保证了纳米材料的高稳定性，而且提高了对H 2S的选择性。HS-SDME法工艺简单，只需一滴溶剂即可完成分析。
有益效果：相对于现有技术，本发明采用智能设备比色法，检测极限为65nM左右，线性范围为0.1μM-100μM，所建立的方法能够应用于测定实际样品如蛋清及牛奶等不透明样本中的H2S，具有流程少、操作简单、检测效率高等优点。
附图说明
图1：利用银-金核壳三角形纳米薄片顶空单滴液相微萃取后检测H 2S方法机理图
图2：制备的Ag TNS和三种不同金层厚度的Ag@Au TNS的TEM图:I(B and C),II(D，E and F)，III(G and H)；其中A为Ag TNS的TEM图；B和C为I层金的Ag@Au TNS的TEM图；D，E和F为II层金的Ag@Au TNS的TEM图；G和H为III层 金的Ag@Au TNS的TEM图。
图3：Ag@Au TNS在100μM H 2S分别孵化5、10、15、20min的TEM图和反应前)后的EDX mapping。其中，1)为Ag@Au TNS在100μM Na 2S分别孵化5、10、15、20min的TEM图；2)为Ag@Au TNS在100μM H 2S反应前的EDX mapping；3)为Ag@Au TNS在100μM H 2S反应后的EDX mapping。
图4：Ag@Au TNS经100μM Na 2S HS-SDME 20分钟后的紫外-可见吸收光谱变化图。
图5：Ag@Au TNS在100μM Na 2S，孵化温度从15℃到35℃；目标物溶液的pH从5到9；孵化时间从0min到20min的优化图。
图6：Ag@Au TNS分别在10mM CO 2、SO 2、NO、HBr、HCl、NH 3和100μM H 2S等挥发性气体中选择性检测图
图7：Ag TNS、Ag@Au TNS I，II，III分别在100μM Na 2S溶液(蓝色)和100μM Na 2SO 3溶液(红色)经HS-SDME 20分钟后吸收光谱的变化图。
图8：H 2S紫外检测的校准曲线。条件：100μM H 2S，pH 6，30℃，孵化20min；
图9：H 2S智能手机纳米比色检测的校准曲线。条件：100μM H 2S，pH 6，30℃，孵化20min；
图10：不同浓度的H 2S对应的紫外光谱图。
图11：牛奶在10天内分别保存在4℃和25℃时硫化氢浓度变化曲线；
图12：鸡蛋在10天内分别保存在4℃和25℃时硫化氢浓度变化曲线。
具体实施方式
以下结合附图和具体实施例，对本发明方法做出进一步说明。
实施例1利用顶空单滴液相微萃取法和智能设备比色测定硫化氢的方法
(1)取50mL烧杯，将40μL硝酸银(0.1M)，600μL柠檬酸钠(0.1M)，112μL双氧水(30％)，加入去离子水至39.6mL，利用磁力搅拌器，在30℃下快速搅拌10min，之后快速加入400μL NaBH 4(0.1M)，同时停止搅拌，此时溶液会形成一个淡黄色的溶液。1-2min后，溶液由黄色逐渐变为红色、绿色、蓝色，说明Ag TNS制备完成。离心后使用去离子水清洗3次，使用前储存在4℃的黑暗中。
(2)将制备好的Ag TNS(20mL)离心洗涤后重新分散于4.5mL去离子水中，对Ag TNS进行横向和纵向的生长，方法如下：加入500μL PVP(乙烯基吡咯单体浓度为 17.5mM)、18.7μL抗坏血酸(0.5M)加入到Ag TNS水溶液中，利用1mL一次性注射器将300μL硝酸银(0.6mM)以0.1mL/min加入到Ag TNS中。随后将150μL柠檬酸钠(0.1M)利用1mL一次性注射器以0.1mL/min加入到Ag TNS中，15min后利用2mL一次性注射器以0.1mL/min将1.5mL硝酸银和柠檬酸钠的混合溶液加入Ag TNS溶液中。溶液混合均匀后，不需进一步纯化，经离心和去离子水清洗后，通过再次加入500μL PVP、75μL二乙胺、100mL抗坏血酸(0.5M)以及含金溶液500μL，使金层沉淀在Ag TNS表面。最后，将产物(Ag@Au TNS)离心，常温下用去离子水洗涤几次，去除AgNO 3沉淀和多余PVP。Ag@Au TNS使用前储存在4℃黑暗中。如图2所示是制备的Ag TNS和三种不同金层厚度的Ag@Au TNS的TEM图。从图2A中可观察到Ag TNS的平均边长约为55nm。Ag@Au TNS的三种不同Au层的厚度分别为0.8nm、1.58nm和2.7nm，如图2B-H所示。
(3)制备样品溶液，由于Na 2S的不稳定性，其水溶液需新鲜配置并在4℃下黑暗中储存。新鲜的牛奶和鸡蛋是直接从超市购买的，没有进一步加工。每个实际样本类型分为两组分别存储在环境温度(25℃)和4℃。如图3所示是Ag@Au TNS萃取H 2S，5、10、15、20min的TEM图，以及Ag@Au TNS萃取前后的EDX mapping.Ag@Au TNS与H 2S接触后虽然边缘萎缩变形，但TNS的总体形状仍保持不变。EDX结果表明，边缘与拐角的元素硫分布无明显差异。
(4)如图1所示，是利用银-金核壳三角形纳米薄片顶空单滴液相微萃取后检测H 2S方法机理图。首先，将1.0mL不同浓度的Na 2S溶液或实际样品加入1.5mL带盖聚丙烯离心管中，然后将3.0μL Ag@Au TNS溶液，滴到顶端盖子内部。借助水的表面张力和分子间作用力，液滴不会轻易从顶部跌落，除非离心管剧烈震动。因此，以这种方式进行SDME是可行的。将盖子快速而轻柔地盖在到离心管上，使离心管静置20分钟，确保H 2S从样品中被释放出来，被萃取液滴充分提取。SDME之后，小心地打开离心管盖，将3.0μL萃取剂使用移液枪转移至NanoPhotometer紫外可见分光光度计分析其紫外可见光信号。对于SNC来说，液滴的照片是由智能手机摄像头拍摄的。该图像的RGB(红、绿、蓝)颜色由EKColorPicker进行分析。图像的R(红色)值直接由EKColorPicker软件提供，用来表示颜色的强度。
加入的PVP、抗坏血酸和柠檬酸钠分次添加均需间隔10min。
硝酸银和柠檬酸钠的混合溶液中的硝酸银浓度为0.75mM，柠檬酸钠浓度为1.13 mM。
所述含金溶液为：400μL PVP(乙烯基吡咯单体浓度为0.5M)，80μL KI(0.2M)，20μL HAuCl 4(0.25M)，3mL超纯水。且以0.05mL/min速度添加。
所述NaBH 4(0.1M)：为冰水制备且现配现用。
(5)数据处理
测取数据：RGB的值由取色软件通过拍摄的图像直接提供，通过测取R(红色)值的数据绘制标准曲线。
绘制标曲：将计算得的R值的差值作为纵坐标，取0.01μM-100μM浓度的对数为横坐标得出标准曲线。
读取数据：通过得到的标准曲线，根据LOD＝3δblack/k，得出最低检测限浓度。其中LOD是指最低检测限，δblack是未反应的Ag@Au TNS的标准偏差，k是标准曲线的斜率。图5为优化实验图，得到最优的实验条件为pH 6，30℃，萃取20min。图8、9分别为H 2S紫外检测的校准曲线和H 2S智能手机纳米比色检测的校准曲线。检测极限分别为7nM和65nM左右，线性范围为10nM-10μM和0.1μM-100μM。
图10为不同浓度的H 2S对应的紫外光谱图。
为评估Ag@Au TNS的选择专一性，对10mM CO 2、SO 2、NO、HBr、HCl、NH 3和100μM H 2S分别萃取研究，如图6所示，对H 2S效果最佳。由于Au层厚度在H 2S的成功检测中起着关键作用，本文还对该参数对灵敏度和选择性的影响进行了评价。在这里，SO 2被用作干扰素。如图7所示，随着Au层厚度的增加，灵敏度略有下降，但选择性增强如图7所示为四种材料在H 2S和SO 2条件下萃取，可得Ag@Au II TNS是最佳的萃取剂。
实施例2牛奶中H 2S的检测实例：
为了检测牛奶中H 2S含量的变化趋势，对鲜牛奶样品中的H 2S进行了测定并连续监测10天。将从超市直接购得的鲜牛奶放置在两个烧杯中进行储存，分别放置在4℃冰箱和室温25℃下保存。使用紫外可见分光光度计测量实验数据，得到牛奶中的H 2S在10天内的变化趋势图，如附图11。为了评估基质效应，这些样品分别加入0.02μM、0.2μM和2μM H 2S，经紫外-可见分光光度计检测；分别加入0.2μM、2μM和20μM、H 2S经智能手机纳米比色法检测。结果分别如表1、2所示，获得了较好的回收率(97-105％)。HS-SDME，完全避免了基质干扰问题，其测取数据和计算方法类似水溶液中的H 2S测 定方式。
表1用HS-SDME-UV-vis法测定新鲜牛奶样品中添加的酸性不稳定硫化物的含量
Figure PCTCN2020077103-appb-000001
a相对回收率＝(总浓度-空白浓度)/掺入浓度
表2用HS-SDME-SNC法测定新鲜牛奶样品中添加的酸性不稳定硫化物的含量
Figure PCTCN2020077103-appb-000002
实施例3鸡蛋中H 2S的检测实例：
为了检测鸡蛋中H 2S含量的变化趋势，对鸡蛋样品中的H 2S进行了测定并连续监测10天。将从超市直接购得的鸡蛋分别放置在4℃冰箱和室温25℃下保存。在鸡蛋壳一端打开直径为5mm的天窗。使用紫外可见分光光度计测量实验数据，得到鸡蛋中的H 2S在10天内的变化趋势图，如附图12。为了评估基质效应，这些样品分别加入0.02μM、0.2μM和2μM H 2S，经紫外-可见分光光度计检测；分别加入0.2μM、2μM和20μM、H 2S经智能手机纳米比色法检测。结果分别如表3、4所示，获得了较好的回收率(95-104％)。
表3用HS-SDME-UV-vis法测定鸡蛋样品中添加的酸性不稳定硫化物的含量
Figure PCTCN2020077103-appb-000003
Figure PCTCN2020077103-appb-000004
表4用HS-SDME-SNC法测定鸡蛋样品中添加的酸性不稳定硫化物的含量
Figure PCTCN2020077103-appb-000005

Claims (8)

  1. 一种利用顶空单滴液相微萃取法和智能设备比色测定硫化氢(H 2S)的方法，其特征在于，包括以下步骤：
    以银-金核壳三角形纳米薄片(Ag@Au TNS)作为纳米检测探针，结合顶空单滴微萃取(HS-SDME)的分析方法，使待测样品中挥发出的H 2S被纳米检测探针特异性萃取，取萃取后的样本，借助智能设备拍照功能和取色软件对H 2S进行检测。
  2. 根据权利要求1所述的测定硫化氢的方法，其特征在于，所述银-金核壳三角形纳米薄片的制备方法包括以下步骤：
    (1)取硝酸银，柠檬酸钠和双氧水加入到去离子水中搅拌，之后快速加入NaBH 4，同时停止搅拌，一定时间后离心，清洗，即得银三角形纳米薄片(Ag TNS)，备用；
    (2)将上述Ag TNS重新分散于去离子水中，加入PVP和抗坏血酸，随后依次滴加硝酸银溶液、柠檬酸钠溶液以及硝酸银和柠檬酸钠的混合溶液，混合均匀后，经离心后用去离子水清洗。
    (3)通过再次加入PVP、二乙胺、抗坏血酸以及含金水溶液，使金层沉淀在Ag TNS表面，最后将产物离心，去离子水洗涤，用于去除AgNO 3沉淀和多余的PVP，即得所述银-金核壳三角形纳米薄片(Ag@Au TNS)。
  3. 根据权利要求2所述的测定硫化氢的方法，其特征在于，步骤(1)中硝酸银，柠檬酸钠和双氧水的摩尔比为1:(10-20):(200-360)，混合溶液与NaBH 4的体积比为(1.3-2.5):1。
  4. 根据权利要求2所述的测定硫化氢的方法，其特征在于，步骤(3)中所述PVP、二乙胺和抗坏血酸的体积比为(1-9):(0.25-1.25):1，含金水溶液的成分包括PVP、KI、HAuCl 4和超纯水，Ag和Au的总物质量之比为(10-4)：1。
  5. 根据权利要求1所述的测定硫化氢的方法，其特征在于，所述纳米检测探针特异性萃取的方法包括以下步骤：
    将待测样品加入带盖容器中，取下盖子，将微量Ag@Au TNS溶液滴到盖子内表面，随后将盖子盖在到容器上，静置一段时间，确保H 2S从样品中释放出来后被萃取液滴充分提取，结束后，打开盖子，其盖内表面液滴即为所述萃取后的样本。
  6. 根据权利要求1所述的测定硫化氢的方法，其特征在于，所述借助智能设备拍照功能和取色软件对H 2S进行检测的方法包括以下步骤：
    测取数据：RGB的值由取色软件通过拍摄的图像直接提供；
    绘制标曲：将计算得的R值的差值作为纵坐标，浓度的对数为横坐标得出标准曲线；
    读取数据：通过得到的标准曲线，根据LOD＝3δblack/k，得出最低检测限浓度；其中LOD是指最低检测限，δblack是空白溶液的标准偏差，k是标准曲线的斜率。
  7. 根据权利要求1所述的测定硫化氢的方法，其特征在于，所述智能设备包括智能手机或平板电脑。
  8. 根据权利要求1所述的测定硫化氢的方法，其特征在于，所述取色软件为EKColorPicker软件、取色器软件或ChemEye。
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