利用雷射光鉗探討膠原蛋白黏彈度受紫外光照射的影響

Using optical tweezers to study the effect on viscoelasticity of collagen by UV irradiation

張世熙(Shih-His Chang)

指導教授 ： 吳見明

國立清華大學/原子科學院/生醫工程與環境科學系/碩士(2012年)

https://doi.org/10.6843/NTHU.2012.00422

摘要

自1970 年，Arthur Ashkin 成功利用雷射光捕捉粒子之後雷射光鉗便不斷創新發展與應用，不僅常用於一個或多個粒子的捕獲和操縱，也可以藉由微流變學的方法用來量測複雜流體的特性。
膠原蛋白是一種天然生醫材料，具有良好的生醫功能特性，可促進軟組織無疤痕形成的修復傷口癒合。然而含膠原蛋白的製品仍須經過滅菌處理，處理過程可能會造成膠原蛋白結構損害，從而降低其生物活性與功能。我們使用波長為254nm紫外光進行滅菌，並由雷射光鉗研究膠原蛋白在滅菌過程黏彈特性的變化，藉由其變化來得知滅菌過程中對於膠原蛋白的損害程度。
首先我們使用蒸餾水作為牛頓流體測試克拉默斯克朗尼希(Kramers–Kronig)轉換式，然後將膠原蛋白配置成濃度0.3(mg/ml)的複合溶液(complex flurds)並加入直徑2μm的聚苯乙烯微球（polystyrene bead），接著使用雷射光鉗捕捉聚苯乙烯微球量測膠原蛋白複雜溶液在254nm紫外光照射下黏彈模數的變化。
量測的結果，樣品溶液皆具有黏彈性的行為，並根據不同的紫外線照射時間改變其黏性及彈性貢獻，其膠原蛋白黏彈度隨著照射紫外光劑量的提高而下降。
本研究使用雷射光鉗與微流變學結合的方法量測膠原蛋白的特性。相對於傳統流變儀的量測方法，使用雷射光鉗與微流變學結合的方法擴大了採樣頻率範圍，也減少了樣品耗材的成本。本實驗量測出膠原蛋白經過照射滅菌劑量後黏彈特性顯著的改變，其彈性模數下降約7倍，黏度下降約5倍。另外我們發現膠原蛋白經過紫外光照射後，黏彈度與凝血功能下降趨勢有相關性。我們推測膠原蛋白可能因為化學鍵被紫外光能量打斷而影響了黏彈特性與凝血功能。本實驗確立了膠原蛋白經紫外光照射滅菌處理過程中，黏彈度下降的趨勢，此結果可以應用在醫學美容或是復健關節炎填充物的研究上。

關鍵字

黏彈模數 ； 膠原蛋白 ； 雷射光鉗 ； 微流變學

並列摘要

Optical tweezers (OT) have been widely applied in various fields ever since Arthur Ashkin successfully pioneered the optical trapping process with laser beam in the 1970s. OT can be utilized to trap and manipulate single or multiple particles, as well as to investigate the mechanical properties of complex fluids through microrheological approach.
Collagen is a natural biomaterial; its biological properties can help prevent scarring during all stages of wound healing process in soft tissues. However, collagen products have to go through sterilization in the processing steps, which can cause structural damage of collagen, and in turns, undermine its bioactive and functional properties. In this paper, optical tweezers are used to study the viscoelasticity of collagen under UV-254 nm irradiation, and to gauge the damaging level of sterilization on collagen.
In the experiment, we first test the Kramers-Kroning procedure using distilled water as Newtonian fluid. Next, 2-μm polystyrene beads are placed in the collagen solution at a polymeric concentration of 0.3 mg/ml. Then, OT are employed to trap and measure the viscoelastic moduli of the collagen polymeric solutions under UV-254 nm irradiation.
Our results indicated that all solutions exhibit viscoelastic behavior. However, different exposure time of UV contributes to the variation in viscous and elastic responses. The viscoelasticity of the collagen goes down as the dosage of UV irradiation rises. Collagen’s elastic modulus and the viscosity decrease about 7 and 5 times, respectively, by sterilization process.
We combined optical tweezers and microrheological technique to measure the viscoelastic properties of collagen solutions. This combination enabled us to extend the range of frequencies and reduced the cost of samples. In the experiment, the viscoelasticity of collagen was measured after a series of irradiations as the sterilization process. We suggested that collagen exposured to UV irradiation may lead to breakage of chemical bonds, thus compromised viscoelasticity and clotting ability. This study concluded the downward viscoelasticity of collagen by UV irradiation in sterilization process. Our results can be used on cosmetology and arthritis rehabilitation.

並列關鍵字

viscoelastic moduli ； collagen ； optical tweezers ； microrheology

參考文獻

61.Wen-Hwar, K., Study on Magnetorheological Damper of Flow Mode. 技術學刊2011年(26卷四期) p. 17-23.

77.葉時孟、洪啟智、陳鴻鈞“尿毒出血之診斷與治療新進展”內科學誌2011年(22期)p.40-47.

1.Ashkin, A., Acceleration and Trapping of Particles by Radiation Pressure.Phys. Rev. Lett., 1970. 24: p. 1-4.

2.James, R.S., Direct observation of the myosin-Va power stroke and its reversal. Nature Structural & Molecular Biology., 2010.17(5): p. 590-595.

3.Wang, M. D., et al., Stretching DNA with optical tweezers. Biophys J., 1997. 72(3): p. 1335-46.