TW201800752A

TW201800752A - Magnetic two dimensional metal nanosheet and method for preparing thereof achieving rapid Raman amplification detecting function and magnetic separation

Info

Publication number: TW201800752A
Application number: TW105119278A
Authority: TW
Inventors: 劉定宇; 陳玨伶; 王冠勛; 詹子儀; 林江珍; 王玉麟; 宮非
Original assignee: 明志科技大學
Priority date: 2016-06-20
Filing date: 2016-06-20
Publication date: 2018-01-01
Also published as: TWI631336B

Abstract

A magnetic two dimensional metal nanosheet, which is prepared by the following steps, comprising step A: synthesizing a carreir with two-dimensional sheet shape; step B: attaching a plurality of metal nanoparticles onto the surface of the carrier with two-dimensional sheet shape; and step C: attaching a plurality of magnetic nanoparticles onto the surface of the carrier with two-dimensional sheet shape. The carrier with two-dimensional sheet shape comprises nano silicon flakes, graphene oxide, or molybdenum disulphide nanosheets; these metal particles are selected from gold nanoparticles or silver nanoparticles; these magnetic nanoparticles comprise iron oxide so that the carrier with two-dimensional shape has magnetic properties. By adding the metal particles and the magnetic nanoparticles to the carrier with two dimensional sheet shape, the magnetic two dimensional metal nanosheet can magnetically separate concentrated materials to be tested and achieve rapid Raman amplification detecting function.

Description

磁性金屬二維奈米片及其製造方法Magnetic metal two-dimensional nano sheet and manufacturing method thereof

本發明係與用於磁分離濃縮及偵測微生物之多功能生物晶片有關；特別是指一種兼具磁性分離及拉曼放大檢測功能之磁性金屬二維奈米片及其製造方法。The present invention relates to a multifunctional biochip for magnetic separation, concentration, and detection of microorganisms; in particular, it relates to a magnetic metal two-dimensional nanosheet with magnetic separation and Raman amplification detection functions and a method for manufacturing the same.

近年由於食品添加物的濫用、工業廢水的排放，以及抗藥型細菌檢測況日費時，對於病菌、水質及食品添加物的檢測需求與技術也日益增加，拉曼散射（raman scattering）是一種利用光散射的原理來測定分子振動（molecular vibration）的光譜學，常被運用於化學、生醫等領域的分析工具。In recent years, due to the abuse of food additives, the discharge of industrial wastewater, and the time-consuming detection of drug-resistant bacteria, the demand and technology for the detection of germs, water quality, and food additives have also increased. Raman scattering is a use The principle of light scattering is used to determine the spectrum of molecular vibration. It is often used as an analytical tool in the fields of chemistry and biomedicine.

拉曼光譜有著以下的優點：Raman spectroscopy has the following advantages:

（1）可用於偵測各種狀態之樣品；(1) Can be used to detect samples in various states;

（2）樣品製備簡易，不需要螢光標定，且不需要長時間培養；(2) sample preparation is simple, no need to use the cursor, and no long-term culture;

（3）可用水當作溶劑，有利於生化分子的檢測。(3) Water can be used as a solvent to facilitate the detection of biochemical molecules.

然而，由於拉曼光譜的樣品前處理需花費大量的時間與人力成本，因此，如何尋找一個較方便且快速的方式達到分離及濃縮待測物的目的，是相關領域人員目前急欲解決的問題。However, because Raman spectroscopy sample preparation requires a lot of time and labor costs, how to find a more convenient and fast way to achieve the purpose of separating and concentrating the analyte is a problem that people in related fields are currently eager to solve. .

有鑑於此，本發明之目的在於提供一種磁性金屬二維奈米片，可快速分離且濃縮待測物，且同時兼具拉曼放大檢測能力，藉此增加拉曼放大檢測技術之方便性及靈敏度。In view of this, the object of the present invention is to provide a magnetic metal two-dimensional nano sheet, which can quickly separate and concentrate the test object, and at the same time has the Raman amplification detection capability, thereby increasing the convenience of Raman amplification detection technology and Sensitivity.

緣以達成上述目的，本發明提供的一種磁性金屬二維奈米片包括有一二維片狀載體、複數個金屬奈米粒子以及複數個磁性奈米粒子。該二維片狀載體係包含一奈米矽片、一氧化石墨烯，或一二硫化鉬奈米片；該些金屬奈米粒子係選自金奈米粒子及銀奈米粒子所構成族群中之至少一者，且附著於該二維片狀載體表面；該些磁性奈米粒子包含一氧化鐵，且附著於該二維片狀載體表面，使該二維片狀載體具有磁性。藉此，該磁性金屬二維奈米片兼具磁性分離濃縮待測物及拉曼放大檢測待測物的功能。In order to achieve the above object, a magnetic metal two-dimensional nano sheet provided by the present invention includes a two-dimensional sheet-shaped carrier, a plurality of metal nano particles, and a plurality of magnetic nano particles. The two-dimensional sheet-shaped carrier system includes a nano silicon wafer, graphene oxide, or a molybdenum disulfide nano wafer; the metal nano particles are selected from the group consisting of gold nano particles and silver nano particles. At least one of them is attached to the surface of the two-dimensional sheet-shaped carrier; the magnetic nano-particles include iron oxide, and is attached to the surface of the two-dimensional sheet-shaped carrier, so that the two-dimensional sheet-shaped carrier is magnetic. Therefore, the magnetic metal two-dimensional nano-sheet has the functions of magnetic separation and concentration of the test object and Raman amplification detection of the test object.

本發明更提供一種磁性金屬二維奈米片的製造方法，包含以下步驟：A、合成一二維片狀載體，其中該二維片狀載體係包含一奈米矽片、一氧化石墨烯，或一二硫化鉬奈米片；B、附著複數個金屬奈米粒子於該二維片狀載體之表面；以及C、附著複數個磁性奈米粒子於該二維片狀載體之表面，使該磁性金屬二維奈米片具有磁性。The invention further provides a method for manufacturing a magnetic metal two-dimensional nano flake, comprising the following steps: A. synthesizing a two-dimensional flake carrier, wherein the two-dimensional flake carrier comprises a nano silicon wafer and graphene oxide, Or a molybdenum disulfide nano sheet; B. attaching a plurality of metallic nano particles to the surface of the two-dimensional sheet-like carrier; and C, attaching a plurality of magnetic nano-particles to the surface of the two-dimensional sheet-like carrier such that the The magnetic metal two-dimensional nano sheet is magnetic.

本發明之效果在於透過該些金屬粒子提高拉曼光譜之檢測訊號，並藉由該些磁性奈米粒子的添加，快速磁分離及濃縮溶液中的待測分子，以此建立一高效能的檢測平台。The effect of the present invention is to increase the detection signal of the Raman spectrum through the metal particles, and to quickly and magnetically separate and concentrate the molecules to be detected in the solution by adding the magnetic nano particles, thereby establishing a highly efficient detection platform.

為能更清楚地說明本發明，茲舉複數實施例並配合圖式詳細說明如後。首先，本案提供之二維奈米片包含一二維片狀載體，以及附著於該二維片狀載體上的複數個金屬粒子或複數個磁性奈米粒子；其中該二維片狀載體包含有奈米矽片（nanoscale silicate platelets，NSP）、氧化石墨烯（graphene oxide，GO）及二硫化鉬（MoS₂ ）奈米片等二維材料；該些金屬粒子則包含金粒子及銀粒子；該些磁性奈米粒子則包含氧化鐵（Fe₃ O₄ ）。以下詳述各實施例之合成步驟。 實施例一：氧化鐵／二維奈米片（磁性二維奈米矽片） 合成磁性二維奈米矽片 In order to explain the present invention more clearly, plural embodiments are described in detail below with reference to the drawings. First, the two-dimensional nano-sheet provided in the present case includes a two-dimensional sheet-like carrier and a plurality of metal particles or magnetic nano-particles attached to the two-dimensional sheet-like carrier; wherein the two-dimensional sheet-like carrier includes Two-dimensional materials such as nanoscale silicate platelets (NSP), graphene oxide (GO), and molybdenum disulfide (MoS ₂ ) nanosheets; these metal particles include gold particles and silver particles; the Some magnetic nano particles contain iron oxide (Fe ₃ O ₄ ). The synthesis steps of the examples are detailed below. Example 1: Synthesis of magnetic two-dimensional nano silicon wafers from iron oxide / two-dimensional nano wafers (magnetic two-dimensional nano silicon wafers)

本實施例之二維片狀載體係包含一奈米矽片10，其中該奈米矽片10的製備方法請見申請人先前專利（中華民國專利公告號I496194–可撓式表面增強拉曼光譜之基板）。首先，將該奈米矽片10透過攪拌分散於25 ml的去離子水中，並進行超聲波處理20分鐘，以獲得良好的分散溶液。接著，將25%的NH₄ OH溶液加入含有奈米矽片10的水溶液中，直到pH值達到11至12。The two-dimensional sheet carrier of this embodiment includes a nano-silicon wafer 10, wherein the preparation method of the nano-silicon wafer 10 is described in the applicant's previous patent (ROC Patent Publication No. I496194-Flexible Surface Enhanced Raman Spectroscopy). Of the substrate). First, the nano silicon wafer 10 was dispersed in 25 ml of deionized water by stirring and subjected to ultrasonic treatment for 20 minutes to obtain a good dispersion solution. Next, a 25% NH ₄ OH solution was added to the aqueous solution containing the nano silicon wafer 10 until the pH reached 11 to 12.

取500mg的氧化鐵（FeSO₄）12，緩慢加入含有奈米薄片10的水溶液中，並持續攪拌12小時；12小時後，將呈灰色的水溶液用去離子水洗滌三次，以去除多餘的NH₄ OH；最後，真空乾燥樣品，以獲得磁性二維奈米矽片100。磁性分離微生物之應用 Take 500mg of iron oxide (FeSO₄) 12 and slowly add it to an aqueous solution containing nanoflakes 10 and continue stirring for 12 hours; after 12 hours, wash the gray aqueous solution three times with deionized water to remove excess NH ₄ OH ; Finally, the sample is vacuum-dried to obtain a magnetic two-dimensional nano silicon wafer 100. Application of magnetic separation of microorganisms

如圖１所示，提供一包含微生物1a之待測溶液1，接著將磁性二維奈米矽片100加入該待測溶液1中（步驟一S1），使該些磁性二維奈米矽片100吸附分布於該微生物1a表面。接著提供一磁鐵2，並將該磁鐵2靠置於該待測溶液1之容器的一側（步驟二S2），使得該些微生物1a被吸引至該容器的該側。藉此，可分離該待測溶液1中的微生物1a，而得到不含有該些微生物1a的分離液1b（步驟三S3）。 實施例二：金奈米粒子／脫層石墨烯二維奈米片 脫層石墨烯二維奈米片 As shown in FIG. 1, a test solution 1 containing microorganisms 1a is provided, and then a magnetic two-dimensional nano silicon wafer 100 is added to the test solution 1 (step S1) to make the magnetic two-dimensional nano silicon wafers. 100 is adsorbed and distributed on the surface of the microorganism 1a. Next, a magnet 2 is provided, and the magnet 2 is placed on one side of the container of the solution 1 to be tested (step S2), so that the microorganisms 1a are attracted to the side of the container. Thereby, the microorganisms 1a in the test solution 1 can be separated, and a separation liquid 1b containing no microorganisms 1a can be obtained (step S3). Example 2: Au Nanoparticles / Delaminated Graphene 2D Nanosheets Delaminated Graphene 2D Nanosheets

本實施例之二維片狀載體係以表面帶正電石墨烯奈米片20c，當然，在其他實施例中，亦可以是奈米矽片或硫化鉬奈米片，而不以石墨烯奈米片為限。以下請參圖２所示，首先進行步驟一（S'1），將未脫層塊狀材料，即片狀石墨20，加入過錳酸鉀（KMnO₄ ）、濃硫酸（H₂ SO₄ ）和濃硝酸（HNO₃ ）的混合液進行氧化反應後，先以過氧化氫水溶液（H₂ O_2(aq) ）洗去反應殘餘離子後，再以去離子水多次洗至溶液接近中性為止，最終，以超音波震盪剝離出脫層氧化石墨烯二維奈米片20a。經上述化學反應後，該片狀石墨20表面將產生許多氧化官能基鍵結，而由於該些氧化官能基具有親水性，故在水溶液中會透過親水作用力以及表面帶負電之靜電排斥力，使得該片狀石墨20能克服層與層間的凡得瓦力以達脫層的效果。合成均勻分散的脫層氧化石墨烯二維奈米片 The two-dimensional sheet carrier of this embodiment is a positively charged graphene nano sheet 20c on the surface. Of course, in other embodiments, it can also be a nano silicon wafer or a molybdenum sulfide nano sheet instead of graphene nano. Rice pieces are limited. Please refer to FIG. 2 as follows. First, step 1 (S'1) is performed, and the delaminated bulk material, namely the flake graphite 20, is added with potassium permanganate (KMnO ₄ ) and concentrated sulfuric acid (H ₂ SO ₄ ). After the oxidation reaction with the mixed solution of concentrated nitric acid (HNO ₃ ), the reaction ions are first washed with an aqueous hydrogen peroxide solution (H ₂ O _{2 (aq)} ), and then washed with deionized water multiple times until the solution is nearly neutral So far, the delaminated graphene oxide two-dimensional nano sheet 20a is finally peeled off by ultrasonic vibration. After the above chemical reaction, the surface of the flake graphite 20 will have many oxidative functional groups bonded, and because these oxidized functional groups are hydrophilic, they will pass through the hydrophilic force and the electrostatic repulsive force negatively charged on the surface in aqueous solution. The flake graphite 20 can overcome the vandal force between layers to achieve the effect of delamination. Synthesis of uniformly dispersed delaminated graphene oxide two-dimensional nanosheets

步驟二（S'2）如下：將脫層氧化石墨烯二維奈米片20a粉末溶於水中，接著將該脫層氧化石墨烯水溶液以超音波震盪約10分鐘，再加入聚乙烯吡咯烷酮（polyvinylpyrrolidone，PVP）後持續攪拌12小時，之後再加入聯氨（N₂ H₄ ）和氨水攪拌一小時，即可得到均勻分散的脫層氧化石墨烯二維奈米片20b。其中聚乙烯吡咯烷酮之結構式為：

合成表面帶正電石墨烯奈米片 Step two (S'2) is as follows: dissolve the delaminated graphene oxide two-dimensional nano flake 20a powder in water, and then sonicate the delaminated graphene oxide aqueous solution for about 10 minutes, and then add polyvinylpyrrolidone (polyvinylpyrrolidone) , PVP), stirring was continued for 12 hours, and then hydrazine (N ₂ H ₄ ) and ammonia water were added and stirred for one hour to obtain a uniformly dispersed delaminated graphene oxide two-dimensional nano sheet 20b. The structural formula of polyvinylpyrrolidone is:

Synthesis of positively charged graphene nano flakes

接著執行步驟三（S'3），將均勻分散的脫層氧化石墨烯二維奈米片20b粉末溶於水中，接著將該溶液以超音波震盪約10分鐘，加入以高分子聚二甲基二烯丙基氯化銨（polydiallyldimethylammoniun chloride，PDDA）22為例的高分子橋接物後，持續攪拌並以90°C加熱迴流12小時，溶液則會逐漸由褐色轉為黑色而成為表面帶正電石墨烯奈米片20c。其中高分子聚二甲基二烯丙基氯化銨22之結構式為：

合成金奈米粒子／石墨烯奈米片 Then perform step three (S'3), dissolve the uniformly dispersed delaminated graphene oxide 2D nano flake 20b powder in water, and then sonicate the solution for about 10 minutes, and then add polymer polydimethyl After the polymer bridge of polydiallyldimethylammoniun chloride (PDDA) 22 as an example, continuous stirring and heating at 90 ° C for 12 hours, the solution will gradually change from brown to black and become positively charged on the surface. Graphene nano sheet 20c. The structural formula of the polymer polydimethyldiallylammonium chloride 22 is:

Synthesis Jinnai Mi particles / graphene nano-sheet

本實施例之金屬粒子為金奈米粒子24，當然，在其他實施例中則不以此為限，例如還可以是銀奈米粒子。步驟四（S'4）首先包含金奈米粒子24的製備，該製備方法係利用檸檬酸鈉熱還原法（citrate thermal reduction method），其中以檸檬酸鈉（Na₃ Ct．2H₂ O）作為還原劑，還原四氯金酸（HAuCl₄ ．3H₂ O）中的金離子（Au³⁺ ），使之還原成原子態的金。詳細實驗步驟如下，將四氯金酸（HAuCl_4(aq) ）加熱至沸騰後再加入檸檬酸鈉（Na₃ Ct_(aq) ），則溶液顏色會由淺黃色逐漸轉為磚紅色，待顏色不再變化，反應即完成。The metal particles in this embodiment are gold nano particles 24, of course, in other embodiments, it is not limited to this, for example, silver nano particles may also be used. Step 4 (S'4) first includes the preparation of gold nano particles 24. This preparation method uses a sodium citrate thermal reduction method, in which sodium citrate (Na ₃ Ct. 2H ₂ O) is used The reducing agent reduces gold ions (Au ³⁺ ) in tetrachloroauric acid (HAuCl _4. 3H ₂ O) to reduce it to atomic gold. The detailed experimental steps are as follows. After heating tetrachloroauric acid (HAuCl _{4 (aq)} ) to boiling and then adding sodium citrate (Na ₃ Ct _(aq) ), the color of the solution will gradually change from light yellow to brick red. No more changes, the reaction is complete.

接著，將含有特定比例之金奈米粒子24的溶液加入含有表面帶正電石墨烯奈米片20c的溶液後，攪拌一小時即可得到含有金奈米粒子／石墨烯奈米片200的水溶液。以X光繞射儀可測得其繞射圖譜，而以穿透式電子顯微鏡則可觀察其微結構。而最終獲得的金奈米粒子／石墨烯奈米片200則可應用於表面增強拉曼光譜之生物分子測試（步驟五，S'5）。 實施例三：磁性金屬二維奈米片 合成磁性金屬二維奈米片 Next, after adding a solution containing a specific ratio of gold nanoparticles 24 to a solution containing a positively charged graphene nanoflake 20c on the surface, stirring for one hour can obtain an aqueous solution containing gold nanoparticles / graphene nanoflake 200. . The diffraction pattern can be measured with an X-ray diffractometer, and the microstructure can be observed with a transmission electron microscope. The final obtained nano-particles / graphene nano-sheets 200 can be applied to the biomolecule testing of surface enhanced Raman spectroscopy (step 5, S'5). Example 3: Synthesis of two-dimensional magnetic metal nanosheets

如圖３所示，本實施例磁性金屬二維奈米片300包含該二維片狀載體，並同時包含該些金屬粒子及該些磁性奈米粒子，其中該二維片狀載體為奈米矽片30（其製備方法請見申請人先前專利：中華民國專利公告號I496194–可撓式表面增強拉曼光譜之基板），而透過穿透式電子顯微鏡下可觀察到其片徑D1約100至1000奈米；該些金屬奈米粒子為金奈米粒子32，其合成於該奈米矽片30上的步驟同樣請見實施例二之S'4，且各該金奈米粒子32的粒徑D2為5-50奈米；該些磁性奈米粒子為氧化鐵（FeSO₄）34，其附著於該奈米矽片30的方式同實施例一，且其粒徑D3為5-500奈米。在其他實施例中，上述磁性金屬二維奈米片的二維片狀載體不以奈米矽片30為限，而也可以是實施例二中的表面帶正電石墨烯奈米片20c（其合成詳見實施例二之S'1至S'3）。磁性分離微生物及生物分子測試之應用 As shown in FIG. 3, the magnetic metal two-dimensional nano sheet 300 of this embodiment includes the two-dimensional sheet-shaped carrier, and also includes the metal particles and the magnetic nano-particles, wherein the two-dimensional sheet-shaped carrier is nano. Silicon wafer 30 (for the preparation method, please refer to the applicant's previous patent: Republic of China Patent Publication No. I496194-a substrate with flexible surface enhanced Raman spectroscopy), and its diameter D1 is about 100 under a transmission electron microscope To 1000 nanometers; the metal nano particles are gold nano particles 32, and the steps for synthesizing them on the nano silicon wafer 30 are also shown in S'4 of the second embodiment. The particle diameter D2 is 5-50 nanometers; the magnetic nanoparticle is iron oxide (FeSO₄) 34, and the method of attaching to the nanometer silicon wafer 30 is the same as that of the first embodiment, and the particle diameter D3 is 5-500 nanometers. Meter. In other embodiments, the two-dimensional sheet-shaped carrier of the magnetic metal two -dimensional nano-sheet is not limited to the nano-silicon wafer 30, but may also be a positively-charged graphene nano-sheet 20 c with a surface in Embodiment 2 ( For the synthesis, see S'1 to S'3 in Example 2). Applied Microbiology and magnetic separation of biological molecules tested

由於該磁性金屬二維奈米片300可透過磁性分離並收集濃縮待測物，例如微生物，亦可利用表面增強拉曼效應（surface-enhanced Raman scattering，SERS）檢測分子物種，其靈敏且專一的訊號得以快速分辨溶液中的汙染物或生物分子。Since the magnetic metal two-dimensional nano-sheet 300 can magnetically separate and collect concentrated analytes, such as microorganisms, it can also detect surface molecular species using surface-enhanced Raman scattering (SERS), which is sensitive and specific The signal allows rapid identification of contaminants or biomolecules in the solution.

以下茲舉該磁性金屬奈米二維奈米片300應用於磁性分離後之SERS檢測實驗結果。不同複合材料於含有腺嘌呤的溶液中所呈現的拉曼光譜 The following is the result of the SERS detection experiment after the magnetic metal nano two-dimensional nano sheet 300 is applied to magnetic separation. Raman spectra of different composite materials in adenine-containing solution

請配合圖４，光譜（a）為使用金屬奈米粒子／二維奈米片所得到的腺嘌呤（adenine，A）拉曼光譜；光譜（b）為使用單獨金屬奈米粒子所得到的腺嘌呤拉曼光譜；光譜（c）為使用金屬奈米粒子／二維奈米片所得到的水溶液（Blank）拉曼光譜；光譜（d）為不添加任何材料下之腺嘌呤的拉曼光譜。其中腺嘌呤於溶液中的濃度為10^-4 M。Please match Figure 4. Spectrum (a) is the adenine (adenine, A) Raman spectrum obtained using metal nano particles / two-dimensional nano tablets; spectrum (b) is the gland obtained using metal nano particles alone. Purine Raman spectrum; spectrum (c) is a Raman spectrum of an aqueous solution (Blank) obtained using metal nano particles / two-dimensional nano flakes; spectrum (d) is a Raman spectrum of adenine without adding any material. The concentration of adenine in the solution is 10 ^-4 M.

觀察使用單獨金屬奈米粒子所得到的腺嘌呤拉曼光譜（光譜b）可知，金屬奈米粒子有很好的拉曼放大能力，相較於對照組腺嘌呤的拉曼光譜（光譜d），放大效果明顯增強。再者，金屬奈米粒子/二維奈米片所得到的腺嘌呤拉曼光譜（光譜a），相較單獨使用金屬奈米粒子所得到的腺嘌呤拉曼光譜（光譜b），則拉曼放大效果更是有3至4倍的顯著增強效果。針對不同化學結構產生特定的光譜 Observing the adenine Raman spectrum (spectrum b) obtained using the metal nanoparticle alone, it can be seen that the metal nanoparticle has a good Raman amplification ability, compared to the Raman spectrum (spectrum d) of the adenine in the control group, The zoom effect is significantly enhanced. In addition, the Raman spectrum (spectrum a) of the adenine obtained by the metal nanoparticle / two-dimensional nanosheet is compared with the Raman spectrum (spectrum b) of the adenine obtained by using the metal nanoparticle alone. The magnification effect has a significant enhancement effect of 3 to 4 times. Generate specific spectra for different chemical structures

如圖５所示，該磁性金屬二維奈米片300可區分組成核酸並帶有編碼遺傳訊息的四種鹼基對：腺嘌呤（A）、胸腺嘧啶（T）、鳥嘌呤（G）及胞嘧啶（C）。測定體內b - 胡蘿蔔素含量 As shown in FIG. 5, the magnetic metal two-dimensional nano-sheet 300 can distinguish four kinds of base pairs constituting a nucleic acid and encoding genetic information: adenine (A), thymine (T), guanine (G), and Cytosine (C). Determination of b - carotene content in the body

b-胡蘿蔔素為一種抗氧化劑，可保護人體免受自由基的侵害，並降低癌症和心臟疾病的發生。而如圖６所示，該磁性金屬奈米複合材料300可用以偵測體內微量的b-胡蘿蔔素的含量，以作為健康指數的指標。水中汙染物體孔雀石綠之定量分析 B-carotene is an antioxidant that protects the body from free radicals and reduces the incidence of cancer and heart disease. As shown in FIG. 6, the magnetic metal nano composite material 300 can be used to detect the trace b-carotene content in the body as an index of a health index. Quantitative analysis of malachite green pollutant in water

近幾年，養殖漁業為殺死水中對魚類有害的菌類而在水中添加孔雀石綠（malachite green），然而其本身具有致癌性，故若孔雀石綠殘留或累積於水產品中，將對人類健康產生潛在性威脅。In recent years, malachite green has been added to water for killing fungi that are harmful to fish. However, it is carcinogenic. Therefore, if malachite green remains or accumulates in aquatic products, it will be harmful to humans. Potential threats to health.

是以，將孔雀石綠作為檢測標的，此實驗利用該磁性金屬奈米複合材料300進行定量分析。觀察圖７之實驗結果可發現，隨著孔雀石綠於水溶液中的濃度提高，拉曼光譜的強度亦隨之提高。Therefore, using malachite green as the detection target, this experiment uses the magnetic metal nanocomposite material 300 for quantitative analysis. Observing the experimental results in Figure 7, it can be found that as the concentration of malachite green in the aqueous solution increases, the intensity of the Raman spectrum also increases.

綜上所述，將本發明該磁性金屬二維奈米片300應用於磁性分離及濃縮技術，可快速前處理待測物，另外，可同時利用表面增強拉曼（SERS）光譜技術以偵測生物分子及水中有害物質之振動光譜，可有效作為一種「光譜指紋」以辨識各分子的種類，進而做到定性及定量的檢測分析。In summary, the magnetic metal two-dimensional nano sheet 300 of the present invention is applied to magnetic separation and concentration technology, which can quickly preprocess the test object. In addition, it can simultaneously use surface enhanced Raman (SERS) spectroscopy technology to detect The vibrational spectrum of biological molecules and harmful substances in water can be effectively used as a "spectral fingerprint" to identify the type of each molecule, and then perform qualitative and quantitative detection and analysis.

另外一提的是，在其他實施例中，該磁性金屬二維奈米片的二維片狀載體並不以奈米矽片或表面帶正電石墨烯奈米片為例，亦可以是二硫化鉬奈米片；該金屬粒子亦可以是銀奈米粒子，而不以金奈米粒子為限；該磁性奈米粒子亦可以其他具磁性的奈米粒子代替氧化鐵。而上述該些實施例同樣可產生上述應用於磁性分離後之SERS檢測的實驗結果。It is also mentioned that, in other embodiments, the two-dimensional sheet carrier of the magnetic metal two-dimensional nano-sheet is not a nano-silicon chip or a positively charged graphene nano-sheet on the surface. Molybdenum sulfide nano flakes; the metal particles may also be silver nano particles, not limited to gold nano particles; the magnetic nano particles may also replace magnetic oxide with other magnetic nano particles. The above-mentioned embodiments can also produce the experimental results of the SERS detection applied to the magnetic separation.

以上所述僅為本發明較佳可行實施例而已，舉凡應用本發明說明書及申請專利範圍所為之等效變化，理應包含在本發明之專利範圍內。The above descriptions are only the preferred and feasible embodiments of the present invention, and any equivalent changes made by applying the description of the present invention and the scope of patent application should be included in the patent scope of the present invention.

［本發明］
1‧‧‧待測溶液
1a‧‧‧微生物
1b‧‧‧分離液
2‧‧‧磁鐵
100‧‧‧磁性二維奈米矽片
10‧‧‧奈米矽片
12‧‧‧氧化鐵
S1‧‧‧步驟一
S2‧‧‧步驟二
S3‧‧‧步驟三
200‧‧‧金奈米粒子／石墨烯奈米片
20‧‧‧片狀石墨
20a‧‧‧脫層氧化石墨烯二維奈米片
20b‧‧‧均勻分散的脫層氧化石墨烯二維奈米片
20c‧‧‧表面帶正電石墨烯奈米片
22‧‧‧高分子聚二甲基二烯丙基氯化銨
24‧‧‧金奈米粒子
S'1‧‧‧步驟一
S'2‧‧‧步驟二
S'3‧‧‧步驟三
S'4‧‧‧步驟四
S'5‧‧‧步驟五
300‧‧‧磁性金屬二維奈米片
30‧‧‧奈米矽片
32‧‧‧金奈米粒子
34‧‧‧氧化鐵
D1‧‧‧片徑
D2‧‧‧粒徑
D3‧‧‧粒徑[this invention]
1‧‧‧ test solution
1a‧‧‧ microorganism
1b‧‧‧ separation liquid
2‧‧‧ magnet
100‧‧‧ Magnetic 2D Nano Silicon Wafer
10‧‧‧Nano silicon wafer
12‧‧‧iron oxide
S1‧‧‧Step 1
S2‧‧‧Step 2
S3‧‧‧Step Three
200‧‧‧gold nano particles / graphene nano flakes
20‧‧‧ flake graphite
20a‧‧‧ Delaminated Graphene Oxide 2D Nanosheets
20b‧‧‧ two-dimensional nanosheets of delaminated graphene oxide uniformly dispersed
20c‧‧‧Positively charged graphene nanosheets
22‧‧‧Polymer polydimethyldiallylammonium chloride
24‧‧‧ Chennai Particles
S'1‧‧‧Step 1
S'2‧‧‧Step 2
S'3‧‧‧Step three
S'4‧‧‧Step 4
S'5‧‧‧Step five
300‧‧‧Magnetic metal two-dimensional nano flakes
30‧‧‧Nano silicon wafer
32‧‧‧Gold Nano Particles
34‧‧‧ Iron oxide
D1‧‧‧ film diameter
D2‧‧‧ particle size
D3‧‧‧ particle size

圖１為本發明第一實施例之使用方法示意圖，揭示氧化鐵／二維奈米片（磁性二維奈米矽片）磁性分離及濃縮溶液中的微生物。圖２為本發明第二實施例之製造方法示意圖，揭示金奈米粒子／脫層石墨烯二維奈米片之合成及用於表面增強拉曼光譜之檢測。圖３為本發明第三實施例之使用方法示意圖，揭示磁性金屬二維奈米片磁性分離微生物並同時進行表面增強拉曼光譜之快速檢測。圖４為拉曼光譜圖，顯示不同複合材料於含有腺嘌呤的溶液中所呈現的拉曼光譜。圖５為拉曼光譜圖，顯示金屬奈米粒子／磁性二維奈米片於不同DNA鹼基對溶液中所呈現的拉曼光譜。圖６為拉曼光譜圖，顯示金屬奈米粒子／磁性二維奈米片於b-胡蘿蔔素溶液中所呈現的拉曼光譜。圖７為拉曼光譜圖，顯示金屬奈米粒子／磁性二維奈米片於不同濃度之孔雀石綠溶液中所呈現的拉曼光譜。FIG. 1 is a schematic diagram of a method for using the first embodiment of the present invention, revealing the magnetic separation and concentration of microorganisms in a magnetic oxide / two-dimensional nano-sheet (magnetic two-dimensional nano-silicon chip). FIG. 2 is a schematic diagram of a manufacturing method according to a second embodiment of the present invention, revealing the synthesis of gold nano particles / delaminated graphene two-dimensional nano flakes and detection of surface enhanced Raman spectrum. FIG. 3 is a schematic diagram of a method of using the third embodiment of the present invention, revealing that magnetic metal two-dimensional nano-sheets magnetically separate microorganisms and perform rapid detection of surface enhanced Raman spectroscopy simultaneously. Figure 4 is a Raman spectrum diagram showing the Raman spectra of different composite materials in adenine-containing solutions. Figure 5 is a Raman spectrum diagram showing the Raman spectra of metal nanoparticle / magnetic two-dimensional nanosheets in different DNA base pair solutions. Figure 6 is a Raman spectrum diagram showing the Raman spectrum of a metal nanoparticle / magnetic two-dimensional nanosheet in a b-carotene solution. Figure 7 is a Raman spectrum diagram showing the Raman spectra of metallic nano-particles / magnetic two-dimensional nano flakes in malachite green solutions of different concentrations.

1a‧‧‧微生物 1a‧‧‧ microorganism

300‧‧‧磁性金屬二維奈米片 300‧‧‧Magnetic metal two-dimensional nano flakes

30‧‧‧奈米矽片 30‧‧‧Nano silicon wafer

32‧‧‧金奈米粒子 32‧‧‧Gold Nano Particles

34‧‧‧氧化鐵 34‧‧‧ Iron oxide

D1‧‧‧片徑 D1‧‧‧ film diameter

D2‧‧‧粒徑 D2‧‧‧ particle size

D3‧‧‧粒徑 D3‧‧‧ particle size

Claims

一種磁性金屬二維奈米片，包含有：一二維片狀載體，係包含一奈米矽片、一氧化石墨烯，或一二硫化鉬奈米片；複數個金屬奈米粒子，選自金奈米粒子及銀奈米粒子所構成族群中之至少一者，且附著於該二維片狀載體表面；以及複數個磁性奈米粒子，包含一氧化鐵，且附著於該二維片狀載體表面，使該二維片狀載體具有磁性；藉此，該磁性金屬二維奈米片兼具磁性分離濃縮待測物及拉曼放大檢測待測物的功能。A magnetic metal two-dimensional nano flake, comprising: a two-dimensional flake carrier comprising a nano silicon wafer, graphene oxide, or a molybdenum disulfide nano flake; a plurality of metal nano particles selected from the group consisting of At least one of the group consisting of gold nanoparticle and silver nanoparticle, and attached to the surface of the two-dimensional sheet-like carrier; and a plurality of magnetic nano-particles, including iron oxide, attached to the two-dimensional sheet-like The surface of the carrier makes the two-dimensional sheet-shaped carrier magnetic; thereby, the magnetic metal two-dimensional nano-sheet has both functions of magnetically separating and concentrating the test object and Raman amplification detection of the test object.

如請求項1所述之磁性金屬二維奈米片，其中該片狀載體更具有複數個高分子聚二甲基二烯丙基氯化銨，使該片狀載體之表面帶有正電。The magnetic metal two-dimensional nano sheet according to claim 1, wherein the sheet-shaped carrier further has a plurality of high molecular polydimethyldiallylammonium chloride, so that the surface of the sheet-shaped carrier is positively charged.

如請求項1所述之磁性金屬二維奈米片，其中該奈米矽片、該氧化石墨烯或該二硫化鉬奈米片之片徑介於100至1000奈米。The magnetic metal two-dimensional nano sheet according to claim 1, wherein a diameter of the nano silicon wafer, the graphene oxide, or the molybdenum disulfide nano sheet is between 100 and 1000 nanometers.

如請求項3所述之磁性金屬二維奈米片，其中各該金屬奈米粒子之粒徑介於5至50奈米。The magnetic metal two-dimensional nano sheet according to claim 3, wherein a particle diameter of each of the metal nano particles is between 5 and 50 nanometers.

如請求項4所述之磁性金屬二維奈米片，其中該些磁性奈米粒子之粒徑介於5至500奈米。The magnetic metal two-dimensional nano sheet according to claim 4, wherein the particle size of the magnetic nano particles is between 5 and 500 nanometers.

一種磁性金屬二維奈米片之製造方法，包含以下步驟： A、合成一二維片狀載體，其中該二維片狀載體係包含一奈米矽片、一氧化石墨烯，或一二硫化鉬奈米片； B、附著複數個金屬奈米粒子於該二維片狀載體之表面；以及 C、附著複數個磁性奈米粒子於該二維片狀載體之表面，使該磁性金屬二維奈米片具有磁性。A method for manufacturing a magnetic metal two-dimensional nano sheet includes the following steps: A. Synthesis of a two-dimensional sheet carrier, wherein the two-dimensional sheet carrier comprises a nano silicon wafer, graphene oxide, or disulfide Molybdenum nano flakes; B. A plurality of metal nano particles are attached to the surface of the two-dimensional flake carrier; and C, a plurality of magnetic nano particles are attached to the surface of the two-dimensional flake carrier to make the magnetic metal two-dimensional Nano flakes are magnetic.

如請求項6所述磁性金屬二維奈米片之製造方法，其中步驟A包含：將一高分子橋接物加入該二維片狀載體中，以合成表面帶正電之二維片狀載體。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 6, wherein step A includes: adding a polymer bridge to the two-dimensional sheet carrier to synthesize a two-dimensional sheet carrier having a positive charge on the surface.

如請求項7所述磁性金屬二維奈米片之製造方法，其中該高分子橋接物包含一高分子聚二甲基二烯丙基氯化銨。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 7, wherein the polymer bridge comprises a polymer polydimethyldiallylammonium chloride.

如請求項6所述磁性金屬二維奈米片之製造方法，其中該二維片狀載體之片徑介於100至1000奈米。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 6, wherein the sheet diameter of the two-dimensional sheet-shaped carrier is between 100 and 1000 nanometers.

如請求項6所述磁性金屬二維奈米片之製造方法，其中步驟B包含：將一含有金屬離子之溶液與一還原劑混和並加熱，使該金屬離子還原產生該些金屬奈米粒子。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 6, wherein step B includes: mixing and heating a solution containing metal ions with a reducing agent to reduce the metal ions to generate the metal nano particles.

如請求項10所述磁性金屬二維奈米片之製造方法，其中該金屬離子包含一金離子，且該含有金離子之溶液包含四氯金酸溶液；該還原劑包含檸檬酸鈉；該些金離子還原後產生該些金奈米粒子。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 10, wherein the metal ion includes a gold ion, and the gold ion-containing solution includes a tetrachloroauric acid solution; the reducing agent includes sodium citrate; The gold nanoparticles are generated after the reduction of gold ions.

如請求項11所述磁性金屬二維奈米片之製造方法，其中各該金奈米粒子之粒徑介於5至50奈米。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 11, wherein a particle diameter of each of the gold nano particles is between 5 and 50 nanometers.

如請求項6所述磁性金屬二維奈米片之製造方法，其中該些磁性奈米粒子包含氧化鐵；於步驟C中，將該二維片狀載體置於pH值介於11至12的環境中，加入該些氧化鐵。The method for manufacturing a magnetic metal two-dimensional nano-sheet as described in claim 6, wherein the magnetic nano-particles include iron oxide; in step C, the two-dimensional sheet-shaped carrier is placed at a pH between 11 and 12. Add these iron oxides to the environment.

如請求項13所述磁性金屬二維奈米片之製造方法，其中該氧化鐵之粒徑介於5至500奈米。The method for manufacturing a magnetic metal two-dimensional nano sheet according to claim 13, wherein the particle size of the iron oxide is between 5 and 500 nanometers.