TWI361082B

TWI361082B - Biocompatible polymer and magnetic nanoparticle with biocompatibilities

Info

Publication number: TWI361082B
Application number: TW97101451A
Authority: TW
Inventors: Wen Hsiang Chang; Wen Uan Hsieh; Shiu Hua Huang; Chin I Lin; Shian Jy Wang; Kelly Teng
Original assignee: Ind Tech Res Inst
Priority date: 2008-01-15
Filing date: 2008-01-15
Publication date: 2012-04-01
Also published as: TW200930409A

Description

1361Q82 九、發明說明：【發明所屬之技術領域】本發明係有關於一種生物相容性高分子，且特別β 關於一種以化學鍵結修飾磁性奈米微粒之生物相容性=分【先前技術】鲁磁振造影（MRI)是多項用於疾病診斷的影像分法之―，屬於非破壞性偵測。歷成像的主要訊號由觀察強磁場下質切旋（magnetie spin)與 =疋率電磁輻射的相關而得。診斷時，將欲分析的器放置於強磁場下，利用無線電波能量的撞擊，接著質子的核磁化產生遲緩現象（relaxation) 而形成影像。 ^ MRI偵測體内影像的偵測極限約在1 mm，這項限制顯不了顯影劑對比度的重要。磁性奈米微粒應用在MRI顯影 _ @上有提升對比度的特性，再使用生物相容性高分子作修 # ’可達到生物相容性效果。若在生物相容性高分子耦接專一性辨識分子及螢光染劑，則此含有專一性辨識分子及營光染劑之磁性奈米微粒可增加偵測專一性、提高靈敏度、及同時觀察螢光訊號。美國專利公開號US2007/0148095 (台灣專利公開號 200724904)中揭示—多功能磁性奈米微粒，並以生物相容性南分子修飾此磁性奈米微粒，且在生物相容性高分子偶合專一性辨識分子及螢光染劑。此生物相容性高分子可 1.361082 5 為：聚乙二醇（PEG)、聚乳酸-聚乙二醇（PLA-PEG)、聚丙交酯（PLA)、聚乙交酯（PGA )、聚己内酯（PCL )、 . 或聚曱基丙稀酸曱醋（PMMA )。美國專利公開號US2006/0216239中揭示一生物相容性高分子用以修飾磁性奈米微粒。市售產品dextrane (Feridex® )或 carboxydextrane ( Resovist® )是主要應用於修飾MRI顯影劑之生物相容性高分子。 • 【發明内容】本發明提供一種生物相容性高分子，其結構如式⑴所示：1361Q82 IX. Description of the Invention: [Technical Field] The present invention relates to a biocompatible polymer, and particularly to a biocompatibility of a magnetic nanoparticle modified by chemical bonding = [Prior Art] Lu magnetic resonance imaging (MRI) is a multi-image classification for disease diagnosis, which is non-destructive detection. The main signal of the calendar image is obtained by observing the correlation between the magnetic spin and the electromagnetic radiation at a high magnetic field. In the diagnosis, the device to be analyzed is placed under a strong magnetic field, and the image is formed by the impact of radio wave energy, followed by proton nuclear magnetization to produce a relaxation. ^ The detection limit of MRI detection in vivo images is about 1 mm. This limitation does not indicate the importance of developer contrast. The use of magnetic nanoparticles in MRI development has improved contrast properties, and biocompatible polymers can be used to achieve biocompatibility. If the biocompatible polymer is coupled to the specific identification molecule and the fluorescent dye, the magnetic nanoparticle containing the specific identification molecule and the camping dye can increase the detection specificity, improve the sensitivity, and simultaneously observe Fluorescent signal. U.S. Patent Publication No. US2007/0148095 (Taiwan Patent Publication No. 200724904) discloses multi-functional magnetic nanoparticles, which are modified with biocompatible south molecules and are specifically compatible with biocompatible polymers. Identify molecules and fluorescent dyes. The biocompatible polymer can be 1.361082 5 as: polyethylene glycol (PEG), polylactic acid-polyethylene glycol (PLA-PEG), polylactide (PLA), polyglycolide (PGA), polyhexyl Lactone (PCL), or polymethyl acrylate vinegar (PMMA). A biocompatible polymer is disclosed in U.S. Patent Publication No. US 2006/0216239 for modifying magnetic nanoparticles. Commercially available products dextrane (Feridex®) or carboxydextrane (Resovist®) are biocompatible polymers that are mainly used to modify MRI developers. • SUMMARY OF THE INVENTION The present invention provides a biocompatible polymer having the structure shown in formula (1):

式(I) R1為具有烷基（alkyl group)、芳香基（aryl gr〇up)、羧基 (carboxyl group)、或胺基（amino gr0Up)至少其一之官能基； R2 為烧基（aikyl gr〇up )、或芳香基（_ gr〇up ); n=5到1000之整數； m=l到1〇之整數。本發明並提供-種具有生物相容性之磁性奈米微粒，包含：一磁性奈米微粒；一以化學鍵結修飾該磁性奈⑽粒之生物相容性高分 1361082Formula (I) R1 is a functional group having at least one of an alkyl group, an aryl gr〇up, a carboxyl group, or an amino gr0Up; R2 is a burning group (aikyl gr) 〇up ) , or an aromatic group (_ gr〇up ); n = an integer from 5 to 1000; m = an integer from 1 to 1. The invention also provides a biocompatible magnetic nanoparticle, comprising: a magnetic nanoparticle; a biocompatible high score of the magnetic nano (10) particle modified by chemical bonding 1361082

子，其結構如式(II)所示：Sub, its structure is shown in formula (II):

(carboxyl group )、或胺基（amino n=5到1000之整數； m=l到10之整數。興P)至少其一之官能基; 貫施例，為讓本發明能更明顯易懂，下文特舉出較佳並配合所附圖式，作詳細說明如下：【實施方式】(carboxyl group), or an amino group (amino n = an integer from 5 to 1000; m = an integer from 1 to 10), at least one of the functional groups; in order to make the invention more understandable, The following is a detailed description and a detailed description of the drawings, as follows: [Embodiment]

本發明提供一種生物相容性高分子，其結構如下所厂、The invention provides a biocompatible polymer, the structure of which is as follows:

Ο 式(I) R1 為具有烷基（alkyl group)、芳香基（aryi gr〇Up)、 (carboxyl group)、或胺基（amino group )至少其一之官基 R2 為烧基（alkyl group )、或芳香基（aryl group ); n=5到1000之整數； 1.361082 . m=l到10之整數。請參閱第1圖，本發明之化合物合成流程係利用丁二酸酐 (succinic anhydride)將聚乙二醇(PEG)末端之氫氧基轉換成為羧基（carboxyl group)，再轉換成為矽烷基（Silane)。其中 R1為具有烷基（alkyl group)、芳香基（aryl group)、羧基 (carboxyl group )、或胺基（amino group )至少其一之官能基， R2可為烧基（alkyl group )、或芳香基（aryi gr0Up)。適合ri 和R2的烧基可以是CrC2〇直鏈或具有分枝結構之烷基。在一 ^ 實施例中1可以是CrQ直鏈或具有分枝結構之烷基，例如甲基，乙基，丙基，異丙基，丁基，第二丁基，特丁基，戊基，異戊基，特戊基，己基，異己基等；R2可以是crC6直鏈或具有分枝結構之烷基，例如甲基，乙基，丙基，異丙基，丁基，第二丁基，特丁基，戊基，異戊基，特戊基，己基，異己基等。適合R1和R2的芳香基可以是C6_Ci2有取代基絲取代基的芳香基，例如笨基，聯苯基，萘基等，且其取代基可擇自：羥 φ 基，鹵烧基，院氧基，氰基，硝基，胺基，或院胺基等。亞曱基（methylene)數目（m)可為i到1〇之整數羥乙烯 (oxyethylene)數目（n)可為5到1〇〇〇 (分子量大約為2〇〇至5_0 g/mole的聚乙二醇）之整數。在一實施例中瓜大約是3 ’ η大約是15。 '經由第1圖流程合成之生物相容性高分子，能將其以化學鍵結方式修飾到氧化鐵奈米微粒表面，可以達到生物相容性修飾之目的。本發明之生物相容性高分子還㈣建立粒子（例如不米粒子师粒子、磁性奈米粒子、或超順磁性粒子） ϋδΖ 與專一性標示分子、榮以酉廢相、或診斷用試劑之間的關係。含：本發月並提供—種具有生物相容性之磁性奈米微粒，包磁性奈米微粒；子，：:如=飾該磁性奈米•之生物相容性高分〇〇R Formula (I) R1 is an alkyl group, an aryl group (aryi gr〇Up), a (carboxyl group), or an amino group, at least one of which is an alkyl group. Or an aryl group; n = an integer from 5 to 1000; 1.361082. m = an integer from 1 to 10. Referring to Fig. 1, the synthesis process of the compound of the present invention converts the hydroxyl group at the terminal of polyethylene glycol (PEG) into a carboxyl group by succinic anhydride, and then converts it into a silane group (Silane). . Wherein R1 is a functional group having at least one of an alkyl group, an aryl group, a carboxyl group, or an amino group, and R2 may be an alkyl group or an aromatic group. Base (aryi gr0Up). The alkyl group suitable for ri and R2 may be a CrC2〇 linear chain or an alkyl group having a branched structure. In one embodiment, 1 may be a straight chain of CrQ or an alkyl group having a branched structure, such as methyl, ethyl, propyl, isopropyl, butyl, t-butyl, tert-butyl, pentyl, Isoamyl, pentyl, hexyl, isohexyl, etc.; R2 may be a straight chain of crC6 or an alkyl group having a branched structure, such as methyl, ethyl, propyl, isopropyl, butyl, second butyl , tert-butyl, pentyl, isopentyl, pentylene, hexyl, isohexyl and the like. The aromatic group suitable for R1 and R2 may be an aromatic group having a C6_Ci2 substituted silk substituent, such as a stupid group, a biphenyl group, a naphthyl group, etc., and the substituent may be selected from the group consisting of: hydroxy φ group, halogen group, courtyard oxygen A group, a cyano group, a nitro group, an amine group, or a hospital amine group. The number of methylene groups (m) may be an integer of i to 1 〇. The number of oxyethylenes (n) may be 5 to 1 Å (Polymer B having a molecular weight of about 2 Å to 5 _0 g/mole) An integer of diol). In one embodiment the melon is approximately 3' η and is approximately 15. The biocompatible polymer synthesized by the process of Figure 1 can be chemically bonded to the surface of iron oxide nanoparticles to achieve biocompatibility modification. The biocompatible polymer of the present invention further (4) establishes particles (for example, non-micro particle particles, magnetic nanoparticles, or superparamagnetic particles) ϋδΖ and specificity-labeling molecules, Rongshen waste phase, or diagnostic reagents. Relationship between. Including: This month and provide a kind of biocompatible magnetic nanoparticle, including magnetic nanoparticle; Sub,:: such as = magnetic nanometer's biocompatibility high score 〇〇

Si：式(Π) R1為具有烧基Ulkyl group)、芳香基（_㈣）、後 (Carb〇xyl gr()up )、或胺基（amin。胸p )至少其一之官能基 n=5到1〇〇〇之整數； m=l到1〇之整數；第2圖為本發明之生物相容性高分子藉由化學鍵社表面的示意圖。磁性奈米微粒之材料較佳為鐵鈷、鎳、或前述之氧化物，亦可為其他任何具有磁性的單鐵料或複合物’其中又以具有超順魏者更佳。此磁性奈_ 以生物相容性高分子修飾後，可進一步將該生物相容性高末端修飾形就躲官能基，例如R1⑽為縣^ group)、或胺基（amin。gr〇up)，以利後續與專一性辨識 f/和螢光染劑進行_，應注意的是此時R1不騎基、口為烷基芳香基無法進行反應以耦接其它分子劑。在-實施例tR1可以賴基（earbGxyl胸 C methylene )蠢日广、此7基夕筑目（m)可為1到10之整數，_ r γ (oxye攀ne)數目（η)可為5到麵（聚乙二醇）之二=二大約是3’η大約是15。其中生物相容“分單層披覆='τ、切粒而形成—核殼結構，其較佳為形成一半^外，貫驗顯林發批冑分子奈綠結射提高磁性太錢粒之r2值，相目前市面上R_ist、Feddex之^^ 倍因此:磁性奈米微粒作為磁振造影(MRI)之顯影劑時，、可進一步提1¾影像對比度。 ^-性辨識分子較佳是以共價鍵與生物相容性高分 :。吊用的專-性辨識分子例如有：抗體、蛋白f、素、_、輕白、核酸、或脂質等。經過上述專-性辨= :耦接後的磁性奈米微粒’其粒徑大約為3_500_孰悉此：根據實際需要選擇專—性賴分子，得到各種標 t奈米錄’以增加其標的效率。·可❹纽分子進行 ^-辨識乳癌細胞上的folate接受器（騰_)，而在此例中， ^酸奸的_方式可细末端具有絲或減的生物相容性南/刀子與之反應形成鍵結。人本發明之雜奈米微粒可叫_螢光染劑，可以提供光子心測上另-種㈣診斷的β，再次確職灶。料染劑較佳亦是以共價鍵與生物相容料分子鍵結。適#$光染劑包Si: Formula (Π) R1 is a group having a burnt group Ulkyl group, an aromatic group (_(tetra)), a rear (Carb〇xyl gr()up), or an amine group (amin. chest p) at least one of the functional groups n=5 An integer of 1 ;; an integer of m = 1 to 1 ;; Fig. 2 is a schematic view of the surface of the biocompatible polymer of the present invention by a chemical bond. The material of the magnetic nanoparticle is preferably iron cobalt, nickel, or the foregoing oxide, and may be any other magnetic single iron or composite, wherein it is more preferable to have super-sweet. After the magnetic nanosphere is modified with a biocompatible polymer, the biocompatible high-end modification can further hide the functional group, for example, R1 (10) is a county group, or an amine group (amin. gr〇up). In order to facilitate the subsequent and specific identification of f / and fluorescent dyes _, it should be noted that at this time R1 does not ride the base, the mouth is an alkyl aryl group can not react to couple other molecular agents. In the embodiment tR1, the base (earbGxyl chest Cmethylene) may be abundance, the 7th base (m) may be an integer of 1 to 10, and the number of _r γ (oxye climbing) (η) may be 5 To the surface (polyethylene glycol) two = two is about 3' η is about 15. Among them, the biocompatible "separated single layer coating = 'τ, granules formed" - core-shell structure, which is preferably formed into half of the outside, the cross-test shows that the forest is issued with a batch of molecular green nano-emissions to improve the magnetic too much grain The r2 value is equivalent to the R_ist and Feddex on the market. Therefore, when the magnetic nanoparticle is used as a developer of magnetic resonance imaging (MRI), the image contrast can be further improved. Valence bond and biocompatibility high score: The specific identification molecules for hanging use are, for example, antibodies, proteins f, hormones, _, light white, nucleic acids, or lipids, etc. After the above-mentioned specificity discrimination = : coupling After the magnetic nanoparticle 'its particle size is about 3_500_ 孰 : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : : 专 : 专Identifying the folate receptor on the breast cancer cells (Teng_), and in this case, the smear of the sorghum can have a silky or reduced biocompatibility South/knife reacting with it to form a bond. The hybrid nanoparticle can be called _fluorescent dye, which can provide photon to the other kind of (4) diagnostic β, and then Indeed functional kitchen. Dye material is also relatively good and covalently bonded molecules biocompatible material. Suitable optical dyes packet # $

<#Μ| V<#Μ| V

、☆ J 1^61082 • 括：有機染劑、無機染劑、或有機金屬錯合物等。螢光染劑的激發與放射光源並無特別限定，可為可見光（VIS)、近紅外光 • ⑽R)、或紫外光（uv) ¥。經過上述螢光染劑與專—性辨識分子耦接後的奈米微粒’其粒徑較佳為15_2〇〇nm。【實施例1 :氡化鐵奈米微粒合成】將 FeCl2 . 4H20 11.6 g，0.058 mole、FeCl3 · 6H2〇 11.6 g, 0.096111〇4與40〇1111去離子水放入三頸瓶中，用攪拌器以3〇〇 _ rPm在25°c環境下攪拌。將2.5 N NaOH ( 170 ml)以固定流速47 ml/hr滴入三頸瓶中，NaOH滴完後反應至測其pH值落於11〜12範圍，加入20 ml油酸於三頸瓶中，使其反應30分鐘。接下來用6 N HC1溶液緩慢調整其pH值約為1，此時包覆油酸之氧化鐵微粒會慢慢沉澱，用去離子水進行水洗4〜5 次’將多餘油酸洗掉，收集被油酸被覆的氧化鐵微粒並抽乾。【實施例2 :生物相容性高分子mPEG-silane合成】 • 將分子量750之甲氧基聚乙二醇(mPEG) 300 g，0.4 mol與 N-曱基 2-四氫0比各酮（N-Methyl-2-Pyrrolidone) 600 ± 5 ml ’ 在1000 ml圓底燒瓶中抽真空（真空度20 Torr) 2小時以上。加入丁二酸酐（succinic anhydride ) 48 g，0.48 mol 與對二曱基胺n比咬（4-Dimethylamino_pyridine , DMAP) 19.5 g，0.159 mol ，在30°C下反應兩天。以1 ml/min的速度加入亞硫醯氯（thionyl chloride) 36 土 1 ml，0.48 mol反應2〜3小時，並排水集氣。接著再以1 ml/min 1361082 * « 的速度加入三乙胺（triethylamine) 133.8ml, 0.96 mol。待溫度冷卻至室溫，過濾移除沉澱物，加入3-丙胺三乙氧基矽烷 (3-Aminopropyl triethoxysilane) 94.5 ml, 0.4 mol 反應至少 8 小時。反應完後滴入9 L異丙趟（isopropyl ether)再沉殿，收集沉殿物加入500 ml曱笨（toluene)回溶，並以5000 rpm離心 5分鐘去除沉澱物。再於上層液滴入9L異丙醚再沉澱。收集棕色油狀液體並在真空下抽乾，得到生物相容性高分子 mPEG-silane ° 【實施例3 :生物相容性高分子COOH-PEG-silane合成】將分子量750之聚乙二醇(PEG) 300 g, 0.4 mol與N-曱基 2-四氫0比各酮（N-Methyl-2-Pyrrolidone) 600 ml，在 1000 ml 圓底燒瓶中抽真空（真空度20 Torr) 2小時以上。加入丁二酸肝（succinic anhydride) 96 ± 1 g，0.96 mol 與對二曱基胺π比咬（4-Dimethylamino-pyridine，DMAP) 39.0 g， 0.318mol ’在30°C下反應兩天，得到兩端具有羧基之聚乙二醇（COOH_PEG )。以1 ml/min的速度加入亞硫醮氣（thionyl chloride) 36ml, 0.48 mol反應2〜3小時，並排水集氣。接著再以1 ml/min的速度加入三乙胺（triethylamine) 133.8 ml, 0.96 mol。待溫度冷卻至室溫，過濾移除沉澱物，加入3-丙胺三乙氧基矽烷 (3-Aminopropyl triethoxysilane) 94.5 ml，0.4 mol 反應至少 8 小時。 12 1361082 ; 反應完後滴入9L異丙醚（isopropylether)再沉澱，收集沉澱物加入500 ml甲笨（t〇iuene)回溶，並以5〇〇〇 rpm離心 5分鐘去除沉澱物。再於上層液滴入9]L異丙醚再沉澱。收集棕色油狀液體並在真空下抽乾，得到生物相容性高分子 COOH-PEG-silane。【實施例4 :氧化鐵奈米微粒表面修飾pEGsilane】取含有10 g氧化鐵之曱苯溶液M 2 L，加入250 g的鲁 mPEG-silane或COOH-PEG-silane，超音波震盪反應2-3小時。於反應溶液中加入1.5 L的去離子水以超過濾濃縮裝置進行純化及濃縮，確定純化完畢後將總體積濃縮至約l〇〇ml，得到具生物相容性高分子修飾之氧化鐵奈米微粒。【實施例5 :專一性辨識分子修飾】取 226 μΐ 葉酸（Folate)溶液(Folate/Dimethyl Sulfoxide: 10 I mg/ml)於 50 ml 掠色圓底瓶中，加入 5 ml Dimethyl Sulfoxide 及 176.5 μΐ Dicyclohexyl Carbodiimide 溶液（Dicyclohexyl Carbodiimide / Dimethyl Sulfoxide: 5 mg/ml)，反應 1 小時後，加入 98.5 μΐ NHS 溶液（N-hydroxysuccinimide/ Dimethyl Sulfoxide: 5 mg/ml)反應1小時，加入289 μΐ乙二胺。此為溶液Α。取經由COOH-PEG-silane修飾後之氧化鐵奈米微粒(IOPC) lml(4.48 mg/ml)置於 50 ml 圓底瓶中，力口入 1〇 ml Dimethyl Sulfoxide，抽真空 1 小時，力π 入 176.5 μΐ Dicyclohexyl 1361082☆ J 1^61082 • Includes: organic dyes, inorganic dyes, or organic metal complexes. The excitation and radiation source of the fluorescent dye are not particularly limited, and may be visible light (VIS), near-infrared light (10) R), or ultraviolet light (uv) ¥. The nanoparticles which have been coupled to the specific-identifying molecules by the above-mentioned fluorescent dye have a particle diameter of preferably 15_2 〇〇 nm. [Example 1: Synthesis of bismuth telluride particles] FeCl2 .4H20 11.6 g, 0.058 mole, FeCl3 · 6H2 〇 11.6 g, 0.096111〇4 and 40〇1111 deionized water were placed in a three-necked flask with a stirrer Stir at 3 ° _ rPm in a 25 ° C environment. 2.5 N NaOH (170 ml) was dropped into a three-necked flask at a fixed flow rate of 47 ml/hr. After the NaOH was dropped, the pH was measured to fall within the range of 11 to 12, and 20 ml of oleic acid was added to the three-necked flask. Allow to react for 30 minutes. Next, slowly adjust the pH value to about 1 with 6 N HC1 solution. At this time, the iron oxide particles coated with oleic acid will precipitate slowly, and wash with deionized water for 4~5 times. 'The excess oleic acid is washed off and collected. The iron oxide particles coated with oleic acid are drained. [Example 2: biocompatible macromolecular mPEG-silane synthesis] • a molecular weight of 750 methoxypolyethylene glycol (mPEG) 300 g, 0.4 mol and N-mercapto 2-tetrahydro 0 ketone ( N-Methyl-2-Pyrrolidone) 600 ± 5 ml ' Vacuum (vacuum 20 Torr) in a 1000 ml round bottom flask for more than 2 hours. Succinic anhydride 48 g, 0.48 mol and 4-dimethylamino-pyridine (DMAP) 19.5 g, 0.159 mol were added and reacted at 30 ° C for two days. Add 1 ml of thionyl chloride 36 soil at a rate of 1 ml/min, react 0.48 mol for 2 to 3 hours, and drain the gas. Triethylamine 133.8 ml, 0.96 mol, was then added at a rate of 1 ml/min 1361082 * «. After the temperature was cooled to room temperature, the precipitate was removed by filtration, and 3-4.5% of 3-aminopropyl triethoxysilane was added, and 0.4 mol was reacted for at least 8 hours. After the reaction, 9 L of isopropyl ether was added dropwise to the temple, and the collected matter was added to 500 ml of toluene for reconstitution, and the precipitate was removed by centrifugation at 5000 rpm for 5 minutes. Then, the upper layer was dropped into 9 L of isopropyl ether and reprecipitated. The brown oily liquid was collected and dried under vacuum to obtain a biocompatible polymer mPEG-silane ° [Example 3: Biocompatible polymer COOH-PEG-silane synthesis] Polyethylene glycol having a molecular weight of 750 ( PEG) 300 g, 0.4 mol with N-Methyl-2-Pyrrolidone 600 ml, vacuum in a 1000 ml round bottom flask (vacuum degree 20 Torr) for more than 2 hours . Add succinic anhydride 96 ± 1 g, 0.96 mol and 4-Dimethylamino-pyridine (DMAP) 39.0 g, 0.318 mol' at 30 ° C for two days. Polyethylene glycol (COOH_PEG) having a carboxyl group at both ends. 36 ml of thionyl chloride was added at a rate of 1 ml/min, and 0.48 mol was reacted for 2 to 3 hours, and the gas was collected by drainage. Then, triethylamine (133.8 ml, 0.96 mol) was added at a rate of 1 ml/min. After the temperature was cooled to room temperature, the precipitate was removed by filtration, and 3-4.5% of 3-aminopropyl triethoxysilane was added, and 0.4 mol was reacted for at least 8 hours. 12 1361082; After the reaction, 9 L of isopropyl ether was added dropwise to reprecipitate, and the precipitate was collected and added to 500 ml of t〇iuene to be dissolved, and centrifuged at 5 rpm for 5 minutes to remove the precipitate. Then, the upper layer was dropped into 9] L isopropyl ether and reprecipitated. The brown oily liquid was collected and dried under vacuum to obtain a biocompatible polymer, COOH-PEG-silane. [Example 4: Surface modification of iron oxide nanoparticles pEGsilane] Take M 2 L of 10 g of iron oxide in benzene, add 250 g of Lum-PEG-silane or COOH-PEG-silane, and supersonic oscillation 2-3 hour. 1.5 L of deionized water was added to the reaction solution to purify and concentrate by ultrafiltration and concentration apparatus. After the purification was completed, the total volume was concentrated to about 10 μml to obtain a biocompatible polymer modified iron oxide nanometer. particle. [Example 5: Specificity identification molecular modification] 226 μL Folate solution (Folate/Dimethyl Sulfoxide: 10 I mg/ml) was placed in a 50 ml fading round bottom flask, and 5 ml of Dimethyl Sulfoxide and 176.5 μM Dicyclohexyl were added. Carbodiimide solution (Dicyclohexyl Carbodiimide / Dimethyl Sulfoxide: 5 mg/ml) was reacted for 1 hour, and then reacted with 98.5 μM NHS solution (N-hydroxysuccinimide/Dimethyl Sulfoxide: 5 mg/ml) for 1 hour, and 289 μM of ethylenediamine was added. This is the solution Α. Take iron oxide nanoparticle (IOPC) modified by COOH-PEG-silane (IOPC) lml (4.48 mg/ml) in a 50 ml round bottom bottle, dilute 1 〇ml Dimethyl Sulfoxide, vacuum for 1 hour, force π Into 176.5 μΐ Dicyclohexyl 1361082

Carbodiimide 溶液（Dicyclohexyl Carbodiimide / Dimethyl Sulfoxide:5 mg/ml)，反應 1 小時後，加入 98·5 μΐ NHS 溶液 (N-hydroxysuccinimide / Dimethyl Sulfoxide:5mg/ml)反應 1 小時。此為溶液B。取溶液A 2895 μΐ (1/2體積），加入溶液B，攪拌反應8小時後，將此溶液裝入透析膜(MWJ000)以水透析。接著再將溶液超過濾濃縮至體積為2 ml，得到有專一性辨識分子修飾之氧化鐵奈米微粒。【實施例6:螢光染劑修飾】取 10_6 mol/ml NIR 染劑 CypHer5E ( Amersham Biosciencese公司）1 m卜加1(T6 mol乙二胺反應1小時，此為溶液A。將2 mg/ml已接枝葉酸之氧化鐵微粒溶於1〇 ml去離子水中’加入 106 mol 1-ethyl-3 (3-dimethylaminopropyl) carbodiimide (EDC)，反應 1 小時後，加入 l〇·6 m〇i NHS (N-hydroxysuccinimide)反應 1 小時。此為溶液 B。將溶液A加入溶液B中授拌反應8小時後，將此溶液裝入透析膜(MW:3000)以水透析。接著再將溶液超過濾濃縮至體積為2 ml’得到有螢光染劑修飾的具有專一性辨識能力之氧化鐵奈米微粒。【實施例7 :弛緩率(relaxivity)測試】 1.361082 經由實施例5修飾後的氧化鐵奈米微粒iTRI_I〇p與 US2006/0216239釗案產品及目前臨床使用的產品Carbodiimide solution (Dicyclohexyl Carbodiimide / Dimethyl Sulfoxide: 5 mg/ml) was reacted for 1 hour, and then added with 98·5 μΐ NHS solution (N-hydroxysuccinimide / Dimethyl Sulfoxide: 5 mg/ml) for 1 hour. This is solution B. Solution A 2895 μΐ (1/2 volume) was taken, solution B was added, and after stirring for 8 hours, the solution was placed in a dialysis membrane (MWJ000) and dialyzed against water. Then, the solution was ultrafiltered and concentrated to a volume of 2 ml to obtain iron oxide nanoparticles having specific identification molecular modification. [Example 6: Fluorescent dye modification] Take 10_6 mol/ml NIR dye CypHer5E (Amersham Biosciencese) 1 m Bujia 1 (T6 mol ethylenediamine reaction for 1 hour, this is solution A. 2 mg/ml The ferric oxide particles which have been grafted with folic acid are dissolved in 1 〇ml of deionized water. '106 mol of 1-ethylaminopropyl carbodiimide (EDC) is added. After 1 hour of reaction, l〇·6 m〇i NHS ( N-hydroxysuccinimide) was reacted for 1 hour. This was solution B. After adding solution A to solution B for 8 hours, the solution was loaded into a dialysis membrane (MW: 3000) and dialyzed against water. To a volume of 2 ml', a specific particle-removing iron oxide nanoparticle having a fluorescent dye modification was obtained. [Example 7: Relaxivity test] 1.361082 Iron oxide nanowire modified by Example 5. Particle iTRI_I〇p and US2006/0216239 products and current clinical products

Resovist'Resovist'

Feridex藉由下列方法比較其弛緩率n、r2值：配製不同濃度氧化鐵奈米微粒溶液(鐵離子濃度〇.卜〇.2、 0.3、0.4、0.5 mM)，以 BRUKER the Minispec mq 20 量測各濃度/谷液之T1或T2遲緩時間(reiaxatj〇ntime)，以遲缓時間倒數為縱軸，溶液濃度為橫軸，求得一線性關係，此線性關係之斜率即為rl或r2弛緩率(reiaxivity)。比較結果如表一所示，顯示經由本發明方法修飾後的氧化鐵奈米微粒其r2值約為目前臨床產品Resovist、Feridex之2 倍，約為US2006/0216239前案產品之1.4倍，具有較高影像之對比度。表一、本發明、前案與目前臨床用2MRI造影劑^與^之比較本發明 US2006/0216239 Resovist Feridex 粒徑 8-12 nm 8-12 nm 4.2 nm 4.8-5.6 nm (TEM) (TEM) (TEM) (TEM) r2 321.8 229 164 160 (mM-s)'1 ±2.3 rl 33.4 ±0.3 23.6 25.4 40 (mM-s)"1 雖然本發明已以較佳實施例揭露如上，然其並非用以 1361082 限定本發明，任何所屬技術領域中具有通常知識者，在不脫離本發明之精神和範圍内，當可作任意之更動與潤飾，因此本發明之保護範圍當視後附之申請專利範圍所界定者為準。 16 1361082 【圖式簡單說明】第1圖為合成流程圖，用以說明本發明之生物相容性高分子的合成流程。第2圖繪示本發明之具有生物相容性高分子修飾之磁性奈米微粒。【主要元件符號說明】無0 17Feridex compares the flaccid rate n and r2 by the following methods: Prepare different concentrations of iron oxide nanoparticle solution (iron ion concentration 〇.2, 0.3, 0.4, 0.5 mM), measured by BRUKER the Minispec mq 20 For each concentration/valley solution, the T1 or T2 lag time (reiaxatj〇ntime), the reciprocal time of the delay time is the vertical axis, and the solution concentration is the horizontal axis, and a linear relationship is obtained. The slope of the linear relationship is the rl or r2 relaxation rate. (reiaxivity). The results of the comparison are shown in Table 1. It shows that the iron oxide nanoparticles modified by the method of the present invention have an r2 value about twice that of the current clinical products Resovist and Feridex, and about 1.4 times that of the US2006/0216239 pre-product. High image contrast. Table 1. Comparison of the present invention, the prior case and the current clinical 2MRI contrast agent ^ and ^ The present invention US2006/0216239 Resovist Feridex particle size 8-12 nm 8-12 nm 4.2 nm 4.8-5.6 nm (TEM) (TEM) ( TEM) (TEM) r2 321.8 229 164 160 (mM-s) '1 ± 2.3 rl 33.4 ± 0.3 23.6 25.4 40 (mM-s) "1 Although the invention has been disclosed above in the preferred embodiment, it is not The invention is defined by the disclosure of the disclosure of the disclosure of the entire disclosure of the entire disclosure of the entire disclosure of the entire disclosure of The definition is final. 16 1361082 [Simple description of the drawings] Fig. 1 is a synthesis flow chart for explaining the synthesis process of the biocompatible polymer of the present invention. Fig. 2 is a view showing the magnetic nanoparticles modified with the biocompatible polymer of the present invention. [Main component symbol description] None 0 17

Claims

1361082 苐 97101451 號 _ 十、申請專利範圍：1361082 苐 97101451 _ X. Patent application scope:

修正日期:100.10.20 1.一種生物相容性高分子，其結構如式⑴所示：式(I) R1為具有烧基（alkyl group )、芳香基（aryl group )、羧基（carboxyl group )、或胺基（amino group )至少其一之官能基； R2 為炫基（alkyl group )、或芳香基（aryl group ); n=5到1000之整數； m=l到10之整數。 2. 如申請專利範圍第1項所述之生物相容性高分子，其中 R1和R2的烷基是Q-Cm直鏈或具有分枝結構之烷基。 3. 如申請專利範圍第1項所述之生物相容性高分子，其中 R1和R2的芳香基是C6-C12有取代基或不具取代基的芳香基。 4. 如申請專利範圍第1項所述之生物相容性高分子，其中 R1為甲基。 5. 如申請專利範圍第1項所述之生物相容性高分子，其中 R2為乙基。 6. 如申請專利範圍第1項所述之生物相容性高分子，其中 η 為 15。 18 1361082 第971014:)1號修正日期:100.10.20 修正本 7. 如申請專利範圍第1項所述之生物相容性高分子，其中 m為3。 8. —種具有生物相容性之磁性奈米微粒，包含：一磁性奈米微粒；一以化學鍵結修飾該磁性奈米微粒之生物相容性高分子，其結構如式(II)所示： .Si： Ri 七。^ 式(II) R1為具有烧基（alkyl group)、芳香基（aryl group)、羧基（carboxyl group )、或胺基（amin〇 gr〇Up )至少其一之官能基； n=5到1000之整數； m=l到10之整數。 9. 如申請專利範圍第8項所述之具有生物相容性之磁性奈米微粒，其中R1為羧基（carb〇xyi group )、或胺基（amin〇 gr〇up) 至少其一之官能基。 10. 如申請專利範圍第8項所述之具有生物相容性之磁性奈米微粒，其中該磁性奈米微粒具有超順磁性。 11. 如申靖專利範圍第8項所述之具有生物相容性之磁性 19 lOOiUOZ 第 97101451 號修正日期:100.】0.20 , 奈米微粒’其中組成該赌奈麵粒之磁性材料包含下列至少其-：鐵、鈷、鎳、或前述之氧化物。 ♦ ^ Θ專则第9項所述之具有生物相容性之磁性不小微粒射於R1接上—專—性辨識分子，該專-性辨識刀子為4几體、番^白暂'、H4* η丄^ 貧白質胜肽、酵素'醣類、醣蛋白、核酸、脂質。 Λ … 13.如申4利_第12項所叙具有生物相容性之磁性奈米微粒，其中該磁性奈米微粒之粒徑為3_5〇〇nm。太、，H_如㈣專·_ 12摘叙具有絲相容性之磁性不求微粒’其t該生物相容性高分子輕接—螢光染劑。太…I5·如申請專利範圍第M項所述之具有生物相容性之磁性奈米微粒，其_有該螢光_與鱗-性_分子之磁性奈米微粒，粒徑為15-200 nm。 16. 如申請專利範圍帛14項所述之具有生物相容性之磁性奈米微粒’其令該螢錢劑《激發或放射光源為可見光、近紅外光、或紫外光。 17. 如申请專利範圍第14項所述之具有生物相容性之磁性不米微粒其令該螢光染劑為有機染劑、無機染劑、或有機金屬錯合物。 ' 18’如申請專利範圍f I4項所述之具有生物相容性之磁性不米Μ粒，其令该螢光染劑與該專一性辨識分子係以共價鍵與 20 1361082 r 第97101451號修正曰期:]00.10·20 修正本該生物相容性高分子鍵結。 19. 如申請專利範圍第14項所述之具有生物相容性之磁性 • 奈米微粒，其t該專一性辨識分子與該生物相容性高分子形成 • -CONH-鍵結。 20. 如申請專利範圍第14項所述之具有生物相容性之磁性奈米微粒，其中該生物相容性高分子係披覆該磁性奈来微粒形成一核殼結構。 • 21.如申請專利範圍第14項所述之具有生物相容性之磁性奈米微粒，其中該生物相容性高分子係形成一單層坡覆於該磁性奈米微粒。Revised: 100.10.20 1. A biocompatible polymer having the structure shown in formula (1): Formula (I) R1 has an alkyl group, an aryl group, a carboxyl group Or an amino group having at least one of its functional groups; R2 is an alkyl group or an aryl group; n = an integer from 5 to 1000; m = an integer from 1 to 10. 2. The biocompatible polymer according to claim 1, wherein the alkyl group of R1 and R2 is a Q-Cm linear or branched alkyl group. 3. The biocompatible polymer according to claim 1, wherein the aryl group of R1 and R2 is a C6-C12 substituted or unsubstituted aryl group. 4. The biocompatible polymer according to claim 1, wherein R1 is a methyl group. 5. The biocompatible polymer according to claim 1, wherein R2 is an ethyl group. 6. The biocompatible polymer according to claim 1, wherein η is 15. 18 1361082 971014:) No. 1 Amendment date: 100.10.20 Amendment 7. The biocompatible polymer of claim 1 wherein m is 3. 8. A biocompatible magnetic nanoparticle comprising: a magnetic nanoparticle; a biocompatible polymer modified by chemical bonding to the magnetic nanoparticle, the structure of which is as shown in formula (II) : .Si: Ri Seven. ^ Formula (II) R1 is a functional group having at least one of an alkyl group, an aryl group, a carboxyl group, or an amine group (amin〇gr〇Up); n=5 to 1000 An integer; m = an integer from 1 to 10. 9. The biocompatible magnetic nanoparticle according to claim 8, wherein R1 is a carboxyl group (carb〇xyi group) or an amine group (amin〇gr〇up) at least one of the functional groups. . 10. The biocompatible magnetic nanoparticle of claim 8, wherein the magnetic nanoparticle has superparamagnetism. 11. Magnetic compatibility of biocompatibility as described in item 8 of the Shenjing patent scope. 19 lOOiUOZ No. 97101451, date of revision: 100.] 0.20, nanoparticles, wherein the magnetic material constituting the gamma surface contains at least the following It is - iron, cobalt, nickel, or an oxide of the foregoing. ♦ ^ ΘSpecial item 9 The biocompatible magnetic non-small particles are shot on R1 - the special identification molecule, the special identification knife is 4 bodies, Fan ^bai temporary', H4* η丄^ Lean white peptide, enzyme 'sugar, glycoprotein, nucleic acid, lipid. Λ ... 13. The biocompatible magnetic nanoparticle as described in claim 4, wherein the magnetic nanoparticle has a particle size of 3_5 〇〇 nm. Too, H_如(四)专· _ 12 to extract the magnetic properties of silk compatibility, do not seek particles ', t the biocompatible polymer light connection - fluorescent dye. Too...I5······················································ Nm. 16. The biocompatible magnetic nanoparticle according to claim 14 of the patent application, wherein the fluorescent agent "excitation or emission source is visible light, near-infrared light, or ultraviolet light. 17. The biocompatible magnetic nanoparticles according to claim 14 of the patent application, wherein the fluorescent dye is an organic dye, an inorganic dye, or an organic metal complex. '18' is a biocompatible magnetic non-rice grain as described in the patent application scope f I4, which causes the fluorescent dye to covalently bond with the specific identification molecule and 20 1361082 r No. 97101451 Corrected flood season:]00.10·20 Corrected the biocompatible polymer bond. 19. The biocompatible magnetic nanoparticle according to claim 14 of the patent application, wherein the specific identification molecule forms a -CONH-bonding with the biocompatible polymer. 20. The biocompatible magnetic nanoparticle according to claim 14, wherein the biocompatible polymer coats the magnetic nanoparticle to form a core-shell structure. 21. The biocompatible magnetic nanoparticle according to claim 14, wherein the biocompatible polymer forms a single layer of the magnetic nanoparticle.