雷射捕捉誘發水中PNIPAM 分子相轉變動態之研究藉由光學顯微技術及時間分析螢光顯微光譜技術

Abstract

聚(N-異丙基丙烯醯胺) (PNIPAM) 是有名的熱反應性聚合物, 當溫度高於臨界溶液溫度(LCST)時會發生相轉變.PNIPAM 的分子結構會因水分子被排除掉而從直鏈狀聚成球狀. 我們認為這個相轉變的過程是一個相當好的生物聚合物,例如:蛋白質的研究範例因其中包含了去除水分子及分子鏈摺疊改變, 因此在過去的幾十年中已經有許多相轉變現象的研究. 然而,直接的相轉變動態的研究仍是相當稀少. 在這項研究中,我們利用雷射捕捉誘發相轉變結合分子光學來研究相轉變動態及時間解析顯微螢光光譜來研究螢光標定的PNIPAM 分子. 藉由結合靜態及時間解析光譜, 我們可以觀測雷射捕捉誘發相轉變及其動態變化. 研究雷射捕捉誘發PNIPAM相轉變於不同溶液中, H2O 及 D2O, 確實透露出不同的相轉變過程. 雷射捕捉會誘發球形粒子於雷射聚焦點. 大小及形成時間與溶劑及雷射強度相當有關以及會隨之改變. 這個結果說明了因溶劑吸收雷射而產生的熱效應. H2O 的吸收係數比D2O 大以致於相轉變可以藉由較小強度的捕捉雷射來成形. 以未標定的PNIPAM 相轉變動態過程為基準,我們觀測了被標定的PNIPAM 分子,VDP-PNIPAM, 的相轉變動態過程. 藉由觀測光譜波長峰值的位移,螢光顯微及顯微光譜上的結果清楚指出了VDP-PNIPAM相轉變動態過程中,分子環境從直鏈狀變為球狀的改變. 除了測量在雷射捕捉誘發相轉變條件下的螢光光譜,我們也在過程中及結束後測量螢光強度衰退. 分析相轉變過程的螢光強度衰退也透露出了分子環境在向轉變過程中的動態改變. 除此之外,我們發現了一個不同於以往的雙色雷射誘發相轉變現象,當我們同時引入紅外光捕捉雷射與一道較弱的紫外光雷射. 當額外的弱紫外光雷射引入時,藉由雷射捕捉所形成的球形粒子會大小上會發生擴張的現象. 在現階段,這項雙色雷射誘發相轉變大小擴張的現象會以兩個說法來解釋, 共振效應及光轉熱效應.

A poly(N-isopropylacrylamide) (PNIPAM) is famous thermo-responsible polymer which shows phase transition above lower critical solution temperature (LCST). PNIPAM collapses into globule from coli state where hydrated water molecules are excluded from the polymer matrix. It is considered that the phase transition behavior including dehydration process and folding change is a good example of biological polymers such as protein, therefore many studies of phase transition behavior of PNIPAM have been examined for more than couple of decades. However direct molecular dynamics of phase transition of PNIPAM is few. In this study, we studied molecular spectroscopic phase transition dynamics of PNIPAM by introducing laser trapping, optical microscopy and time-resolved fluorescence microspectroscopy with fluorescently labeled PNIPAM. We induce local phase transition around the focal spot of trapping laser by tight focusing of trapping light source under microscope. Laser-induced phase transitin behavior and its dynamics was visualized by combining with steady state and time-resolved spectroscopy. Studies on a trapping-induced phase transition behavior of PNIPAM in different aqueous solvents, H2O and D2O, revealed that clearly different phase transition behavior. Trapping laser irradiation induced spherical particle formation at the focal spot. The size and formation time strongly depends and changes upon solvent and laser power. It is interpreted due to a thermal effect caused by an absorption of irradiated trapping laser light by the solvents. Larger absorption coefficient of H2O than D2O enables the phase transition at lower laser power in former solution. Based on the information of phase transition dynamics obtained by non-labeled PNIPAM, we examined phase transition dynamics of fluorescently labeled VDP-PNIPAM. Fluorescence microscopy and microspectroscopy results of phase transition dynamics of VDP-PNIPAM clearly indicate the molecular environmental change from coli to globule during phase transition with showing the peak wavelength shift of fluorescence spectra. In addition to the fluorescence spectrum under laser trapping-induced phase transition, we measured fluorescence decay during and after phase transition. Analysis of fluorescence decay curves showed the change of decay component during phase transition which also reveal dynamic change of molecular environment during phase transition. Additionally we found an unusual two color lasers induced phase transition behavior when we introduce weak ultraviolet (UV) laser simultaneously with near infrared (NIR) trapping laser. A spherical particle formed by trapping laser irradiation

showed an expansion of its size when an additional weak UV light was introduced. Observed two-color laser irradiation-induced particle size expansion will be interpreted by two reasons at the moment; resonance effect and photothermal effect.