New fluorescent optical pH sensors with minimal effects of ionic strength

摘要

The thesis describes the development, characterization and application of fluorescence-based, optical pH sensors. Special attention is given to the dependence of the sensor signal and changes of ionic strength in the analyte solution. Based on three different methods for minimization of this dependence, various sensor membranes are presented in detail. Further, a new concept to reference fluorescence intensity signals is introduced and applied to an optical pH sensor.Chapter 1 emphasizes the necessity of precise pH control and measurements by means of examples. An overview of possible fields of pH sensors is given in general. In particular, three different formats of optical pH sensors are presented. Furthermore, the cross-sensitivity of the calibration curve of optical sensors towards ionic strength is mentioned.At the beginning of chapter 2, a short, review on the development of the term �pH� is given, followed by the explanation of the principle of optical pH sensors. The effect of ionic strength on the signal of optical pH sensor is explained by means of the law of Debye and Hückel and the definition of activity coefficients. A paragraph about the state of the art in optical pH sensor technology is followed by the second half of chapter 2, concerning the phenomena luminescence. Beside from basics, also methods for referencing and measurement techniques are explained.Chapter 3 describes the application and spectral properties of commercially available, pH-sensitive fluorescent dyes. HPTS, carboxyfluorescein and fluorescein were checked on their cross-sensitivity towards IS in the range from 25 to 500 mM. According to the theory of Debye and Hückel, the two-fold negative charged indicator fluorescein is less affected by IS than HPTS which carries four negative charges. A novel, partially positive charged indicator shows a contrary change of the dissociation constant. In an equimolar mixture with carboxyfluorescein, the effect of IS was distinctly reduced. In chapter 4, two methods are presented based on the principle described previously for the compensation of the effect of IS. The pH-indicator carboxyfluorescein was immobilized on partially amino-modified carboxycellulose. For the first method, the remaining carboxy groups were converted to positively charged groups. Again, mixing positively and negatively charged celluloses made an improvement of the cross-sensitivity towards IS in the range from 25 mM to 500 mM possible. For the second method, the negatively charged cellulose strand was partially loaded with negative charges. Six differently charged sensors were checked on their cross-sensitivity towards IS. One sensor shows a minimal cross-sensitivity towards IS and it shows the smallest zeta-potential, meaning a low charge density and a successful compensation of negative and positive charges. Chapter 5 deals with the third method for minimization the cross-sensitivity towards IS. Novel, fluorescein-based, lipophilic pH-indicators were embedded in an ion-permeable, charge-free polymer. The indicators were made lipophilic by esterification of the carboxy group with a C18 alkyl chain. This ester-modification reduces the number of charges to one and zero for basic and acid form, respectively. As a result of the charge reduction, the effect of IS is reduced to minimum and becomes negligible in the range from 25 to 500 mM. The fluorescent dyes differ in their substituents at 2�- and 7�-position of the xanthene structure. This variation of substituents results in dissociation constants between 5.5 and 8.5. Sensor properties like photo stability, temperature dependence and fluorescent lifetime were analyzed and discussed in detail. Finally, two applications using these sensor membranes were demonstrated.The chapter 6 deals with new amino-modified polymers. The polymers were embedded in hydrogel together with pH-inert reference particles. The fluorescence intensity of the sensors is converted into a phase shift by means of a novel referencing method (Dual Lifetime Referencing) using luminophores with different fluorescent decay times.