OPTICAL SENSOR SYSTEM FOR pH DETERMINATION INDEPENDENTLY OF THE ION STRENGTH USING FLUORESCEIN BOUND TO A POLYMER VIA A URETHANE AND/OR UREA GROUP

Abstract: The invention relates to an optical sensor for pH determination independent of the ion strength which comprises a solid polyurethane, polyurea-urethane or polyurea composition containing pH-sensitive fluorescein bound via a urea or urethane group in the form of a membrane on a transparent support material. The invention also relates to an optical fluorescence method which permits high-accuracy pH determination independent of the ion strength of the measurement solution. The method is particularly suitable for determining the pH of physiological solutions, in particular for determining the pH of blood.

Description: Optical sensor system for pH determination independently of the ion strength using fluorescem bound to a polymer via a urethane and/or urea group The invention relates to an optical sensor for pH determination independently of the ion strength which comprises a solid polyurethane, polyurea-urethane or polyurea composition containing pH-sensitive fluorescem bound via a urethane and/or urea group in the form of a membrane on a transparent support material The invention also relates to an optical fluorescence method which permits high-accuracy pH determination independently of the ion strength of the measurement solution The method is particularly suitable for determining the pH of physiological solutions, in particular for determining the pH of blood It is known that the pK a of an indicator changes with the ion strength of a solution and this change is dependent on the magnitude of the charge on the indicator Thus it has already been proposed in DE-A-3 430 935 to calculate the ion strength and the pH from the difference between the measurement values of two sensors which are dependent on the ton strength to different extents, of which one has the lowest possible dependence on ion strength after calibration with known measurement solutions The sensor described therein which is virtually independent of the ion strength is not precisely in the physiological pH range and has low resolution These sensors are constructed without embedding in a polymer matrix and thus have the disadvantage that the dye is in direct contact with the measurement solution The same fluorescent dye in each of the sensors is immobilized directly on the surface of glass supports via bridging groups, the first sensor additionally containing charges for achieving high polarity and ion-strength dependence, and the other sensor being modified in such a way that it is essentially nonpolar, hydrophobic and independent of the ion strength A major disadvantage of these sensors is that the fluorescence dye is directly affected by the measurement solutions, and both physical influences (for example dissolution of the dye, deposition on the surface) and chemical influences (decomposition of the dye) quickly render the sensors unusable In addition, on excitation in an evanescent field, interference between the evanescent measurement field and fluorescence of the measurement sample cannot be avoided completely, which reduces the measurement accuracy On the other hand, the response time of these sensors is short, since the fluorescent dye bound to the surface comes immediately into contact with the measurement solution The sensitivity is regarded as adequate The optical pH determination method using two sensors which respond to different degrees to the ion strength of a measurement solution requires complex equipment, and a subsequent additional calculation step must be carried out WO 90/00572 describes a sensor for pH determination and partial pressure determination of CO z which comprises a polyurethane hydrogel in which up to 20 % of an azo dye has been copolymerized The azo dye changes its absorption spectrum as a function of the pH, so that pH determination can be carried out via an absorption measurement The effect of the ion strength on the measurement result is not described EP-A-481 740 describes in general terms polyether-polyurethanes in combination with a covalently bound fluorescence dye for pH determination, but no mention is made anywhere of the extent of the effect of ion strength on the measured signal WO 94/28786 describes, for example, a composition of polyurethane acrylate prepolymer and fluorescem acrylamide which is copolymerized on a glass fibre with the aid of a photoinitiator, giving a pH-sensitive sensor whose sensitivity range can be varied within narrow limits by the prepolymer fluorescem acrylamide ratio Nothing is known on the dependence of the measurement result on the ion strength of the solution being investigated It has now been found that selection of very particular polyurethane compositions in combination with a fluorescem dye which is bound to the polymer via a bridging group and a urea group or directly via a urea group allows the production of an optical pH sensor which permits optical pH measurement of high accuracy in the physiological pH range from 6 5 to 8 2 independently of the ion strength, making a second measurement and the calculation step for eliminating the ion strength superfluous It is essential here that the fluorescent dye is bonded to the polymer in only small amounts High accuracy of pH measurement is of particularly great importance in the analysis of human blood, since this measurement can be used, for example, for therapy control of metabolic disorders It is therefore particularly advantageous for rapid and inexpensive investigation if only a single sensor need be employed for this purpose This also allows the analytical instruments to be miniaturized more easily The service life of these sensors is long, since the polymer matrix effectively protects the fluorescent dye against harmful or interfering effects of the measurement medium This does not reduce the sensitivity, and the response times are surprisingly short The polymer compositions allow, for example, the hydrophilicity, hydrophobicity, polarity and/or dielectric constant of the matrix to be set very precisely, which, in combination with the fluorophore concentration range selected, results in measurement independent of ion strength in a certain pH range In spite of embedding of the fluorophore, the response times and conditioning times correspond to the short periods required for optical measurement systems, these parameters being essentially dependent on the membrane thickness For example a response time of about 10 seconds can be achieved by means of the novel sensors The invention relates to an optical sensor for pH determination independently of the ion strength in the physiological range, comprising A) an optically transparent support material, B) a plasticizer-free, thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea which is soluble in organic solvents, in the form of a solid membrane formed from a) 10-40 % by weight of an aromatic, cycloaliphatic or linear aliphatic dnsocyanate b) 50-85 % by weight of a polyethylene glycol having a molecular weight of 2,000-10,000 daltons, c) 0-30 % by weight of a polytetrahydrofuran, ammopropyl-terminated polytetrahydrofuran, polypropylene glycol or ammopropyl-terminated polypropylene glycol having a molecular weight of 600-10,000 daltons, d) 0-10 % by weight of a linear or branched C 2 -C-| 2 alkylenedιol or C 2 -C 12 alkylenedιamιne, e) 0-2 % by weight of a linear or branched C 3 -C 12 alkylenetrιol and f) 0 1-3 % by weight of a fluorescem dye containing an ammo or hydroxyl group bonded directly or at the end of a bridging group, where the percentages are based on the amount of polymer, and the amounts a) to f) total 100 The geometrical form of the support material and thus of the sensor can vary greatly, it can be, for example, a fibre, cylinder, sphere, rectangular block or cube Through-flow systems in which continuous measurements or successive measurements can be carried out are also possible Planar sensors are preferred They can have an area of from 0 01 to about 50 cm 2 , advantageously from 0 02 to 10 cm 2 The measurement region of the sensor can have an area of less than 5 mm 2 , preferably less than or equal to 2 mm 2 The measurement region may be identical with a fully coated surface of the support material Coating of the support material on both sides, but in a locally separated manner, is advantageous The sensor can comprise one or more locally separated membrane layers; in the latter case, parallel measurements with identical or different measurement samples can be carried out The support material is transparent It can be, for example, an inorganic glass, quartz or a transparent thermoplastic or crossimked plastic, such as polycarbonate, polyester, polyamide, polyacrylate or polymethacrylate The dnsocyanates are preferably selected from the group consisting of 1 ,6-bιsιsocyanato- hexane, 5-ιsocyanato-3-(ιsocyanatomethyl)-1,1 ,3-tπmethylcyclohexane, 1 ,3-bιs[5-ιso- cyanato-1 ,3,3-trιmethylphenyl]-2,4-dιoxo-1 ,3-dιazetιdιne, 3,6-bιs[9-ιsocyanatononyl]-4,5-dι- ( 1 -heptenyl)cyclohexene, bιs[4-ιsocyanatocyclohexyl]methane, trans-1 ,4-bιsιsocyanato- cyclohexane, 1 ,3-bιs[ιsocyanatomethyl]benzene, 1 ,3-bιs[1-ιsocyanato-1- methylethyl]benzene, 1 ,4-bιs[2-ιsocyanatoethyl]cyclohexane, 1 ,3- bisisocyanatomethylcyclohexane 1 ,4-bιs[1-ιsocyanato-1-methylethyl]benzene, bιs[ιsocyanatojιsododecylbenzene, 1 ,4-bιsιsocyanatobenzene, 2,4-bιsιsocyanatotoluene, 2,6-bιsιsocyanatotoluene, 2,4-/2,6-bιsιsocyanatotoluene, N,N'-bιs[3-ιsocyanato-4- methylphenyl]urea, 1 ,4-bιs[3-ιsocyanato-4-methylphenyl]-2,4-dιoxo-1 ,3-dιazetιdιne, 1 ,3-bιs[3- ιsocyanato-4-methylphenyl]-2,4,5-tπoxoιmιdazolιdιne, bιs[2-ιsocyanatophenyl]methane, (2- isocyanatophenyl) (4-ιsocyanatophenyl)methane, bιs[4-ιsocyanatophenyl]methane, 1 ,5- bisisocyanatonaphthalene or 4,4'-bιsιsocyanato-3,3'-dιmethyl-bιphenyl The dnsocyanate is particularly preferably bιs[4-ιsocyanatophenyl]methane (4,4'-MDI), 2,4- or 2,6-bιsιsocyanatotoluene (TDI), 1 ,6-bιsιsocyanatohexane (HDI), 5-ιsocyanato-3-ιso- cyanatomethyl-1 ,1 3-trιmethylcyclohexane (IPDI) or bιs[4-ιsocyanatocyclohexyl]methane (MDI), or a mixture of these dnsocyanates Examples of C 2 -C 12 alkylenediols are ethylene glycol, 1 ,3-propanediol, 1 ,4-butanedιol, 1 ,5-pentanedιol, 1 ,6-hexanediol, 1 ,7-heptanediol, 1 ,8-octanediol, 1 ,9-nonanediol, 1 ,10-decanediol, 1 ,11-undecanediol and 1,12-dodecanediol. The diol, component d), in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is preferably ethylene glycol, butanediol or hexanediol. Examples of C 2 -C 12 alkylenediamines are 1 ,2-ethylenediamine, 1,3-dιaminopropane, 1 ,4-diaminobutane, 1 ,5-diaminopentane, 1 ,6-diaminohexane, 1 ,7-diaminoheptane, 1 ,8-diaminooctane, 1 ,9-diaminononane, 1 ,10-diaminodecane, 1 ,11-diaminoundecane and 1 , 12-diaminododecane. Examples of C 3 -C 12 alkylenetriols are glycerol, the various positional isomers of butanetriol, pentanetriol, hexanetriol, heptanetriol, octanetriol, nonanetriol, decanetriol, undecanetriol and dodecanetriol. Preference is given to 1 ,1 ,1-tris{hydroxymethyl)ethane. In the solid membrane of the sensor, the amino or hydroxyl group of the fluorescein dye f) and the terminal isocyanate component a) are bonded in a urea or urethane group. Component b) in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is preferably 70-85 % by weight of polyethylene glycol having a molecular weight of 600-2,000 daltons. Component f) in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is preferably 0.1-1 % of a fluorescein dye which is amino- or hydroxyl- functionalized via a bridging group. The thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea preferably has a molecular weight of from 10,000 to 30,000 daltons. The thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea preferably includes a fluorescein of the formula (la) or (lb) in which F^ is a bridging group and is linear or branched C 1 -C 2 oalkylene, C 3 -C 2 oalkenylene, C 3 -C 20 alkynylene or CH 2 -(O-(CHR 2 ) r -CH 2 -CH 2 )n, R 2 is C 1 -C 4 alkyl, X is OH or NH 2 , r is 0 or 1 , and n is a number from 1 to 20 Another preferred fluorescein is 4-amιnofluoresceιn. Examples of linear or branched C r C 2 oalkyl radicals are methyl, ethyl, and the various positional isomers of propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, Tndecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl and eicosyl Examples of linear or branched C 3 -C 20 alkenyl radicals are the various positional isomers of propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl, undecenyl, dodecenyl, tπdecenyl, tetradecenyl, pentadecenyl, hexadecenyl, heptadecenyl, octadecenyl, nonadecenyl and eicosenyl. Examples of linear or branched C 3 -C 2 oalkynyl radicals are the various positional isomers of propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl, undecynyl, dodecynyl, tndecynyl, tetradecynyl, pentadecynyl, hexadecynyl, heptadecynyl, octadecynyl, nonadecynyl and eicosynyl The thickness of the polymer layer B) is preferably from 0.1 to 500 μm, particularly preferably 1 to 100 μm These layers can be produced in a manner known per se, for example by dissolving the composition in an organic solvent, then casting the solution to give a film, and finally removing the solvent It is also conceivable to produce the layers by methods as known from surface-coating technology Examples are spin-coating, spraying or knife coating, preference being given to spin-coating and spraying Suitable solvents are alcohols, ethers, esters, acid amides and ketones Relatively volatile solvents, in particular tetrahydrofuran, are particularly suitable Besides these methods in which the composition is firstly dissolved then shaped, and the solvent is then evaporated again, hot shaping processes are also possible, since the composition is a thermoplastic material. Suitable processes are extrusion, injection moulding, compression moulding or blow moulding, as known from thermoplastic processing The layer can be transparent or slightly opaque It is preferably transparent In order to improve adhesion, the support materials can be treated in advance with adhesion promoters For the same purpose, plasma treatment of the support material in order to generate functional groups on the surface is also possible The surface can also be provided with copolymeπzable groups in order to achieve particularly high adhesion Examples of known adhesion promoters for glasses are tnethoxyglycidyloxysilane, 3-azιdopropyltπethoxy- silane and 3-amιnopropyltrιethoxysιlane The surfaces treated in this way can be modified further, for example by means of 0-(N-succmιmιdyl)-6-(4'-azιdo-2'-nιtrophenylamιno)- hexanoate It has been found particularly advantageous to treat the surfaces with silanes of ethylenically unsaturated carboxylic acid esters, for example 3-tπmethoxysιlylpropyl methacrylate The polymers can be prepared by methods known per se The reactants can be reacted with one another as such, i e without addition of a solvent or diluent, for example in the melt However, the addition of a solvent or diluent or a mixture of solvents is usually advantageous Examples of such solvents or diluents which may be mentioned are esters, such as ethyl acetate, ethers, such as diethyl ether, dipropyl ether, dnsopropyl ether, dibutyl ether, tert-butyl methyl ether, tetrahydrofuran or dioxan, ketones, such as acetone, methyl ethyl ketone or methyl isobutyl ketone, amides, such as N,N-dι- methylformamide, N,N-dιethylformamιde, N,N-dιmethylacetamιde, N-methylpyrrolidone or hexamethylphosphoπc tnamide, nitπles, such as acetonitπle or propionitπle, and sulphoxides such as dimethyl sulphoxide The amine- or hydroxyl-functional fluoresceins of the formula (la) or (lb) covalently bonded to the isocyanate group can be prepared by processes known per se; the starting materials are either commercial products or can be prepared by analogous processes The polymer-bound fluorescem dyes have strong absorption and fluorescence In principle, both properties can be used for pH measurement However, it has generally been found more favourable and sensitive to carry out fluorescence measurement The invention furthermore relates to a method for the reversible, optical determination of the pH of an aqueous sample independently of the ion strength by the fluorescence method, in which an optical sensor comprising A) a transparent support material, B) a plasticizer-free, thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea which is soluble in organic solvents, formed from a) 10-40 % by weight of an aromatic, cycloaliphatic or linear aliphatic dusocyanate, b) 50-85 % by weight of a polyethylene glycol having a molecular weight of 2,000-10 000 daltons, c) 0-30 % by weight of a polytetrahydrofuran, ammopropyl-terminated polytetrahydrofuran, polypropylene glycol or ammopropyl-terminated polypropylene glycol having a molecular weight of 600-10,000 daltons, d) 0-10 % by weight of a linear or branched C 2 -C 12 alkylenedιol or C 2 -C 12 alkylenedιamιne, e) 0-2 % by weight of a linear or branched C 3 -C 12 alkylenetrιol and f) 0 1-3 % by weight of a fluorescem dye containing an ammo or hydroxyl group bonded directly or at the end of a bridging group, where the percentages are based on the amount of polymer, and the amounts a) to f) total 100, is brought into contact with an aqueous measurement sample and exposed to excitation light, the fluorescence is measured, and the pH is calculated from the measured fluorescence intensity taking into account calibration curves The preferences described above for the polymers and fluorescem dyes apply correspondingly to the method In detail, the method can be carried out by, after calibration with samples of known pH, measuring the fluorescence intensity in contact with a measurement solution of unknown composition, and determining the pH for the measured fluorescence intensity directly from the calibration The sensors are brought into contact with the calibration solutions or with the measurement samples This can be done by hand (for example by pipetting) or by means of a suitable automatic through-flow system, with the sensors permanently installed in a flow cell Such through-flow cells are known to the person skilled in the art and can easily be adapted to the particular application Light sources which can be used for the fluorescence excitation are UV lamps (for example mercury vapour lamps and halogen lamps), lasers, diode lasers and light diodes It may be expedient to use filters to filter out light having the wavelength at which the fluorescent dye has an absorption maximum The fluorescent light emitted by the sensors can be collected, for example using a lens system, and then directed to a detector, for example a secondary electron multiplier or a photodiode The lens system can be arranged in such a way that the fluorescence radiation is measured through the transparent support, over the edges of the support or via the analyte sample The radiation is advantageously directed in a manner known per se via a dichroic mirror The fluorescence from the sensors is preferably measured during contact with the calibration or sample solutions The measurement can be carried out under photostationary conditions with continuous exposure, or, if required, with time resolution This can be achieved, for example, by means of a time-limited laser pulse or by modulation of the intensity of the light source The measurement solution preferably has a pH of from 6 7 to 7.8 The measurement solution can include salts of inorganic or organic acids Examples are salts of citric acid, tactic acid, acetic acid, phosphoric acid, hydrochloric acid or sulphuric The ion strength of the measurement solution is preferably from 0 05 to 5 mol/l, particularly preferably from 0 05 to 1 mol/l The ion strength is preferably built up essentially by 1 1 or 1 2 salts Examples of 1 1 salts are LiCI, NaCl, KCI and NH 4 CI Examples of 1 2 salts are CaCI 2 , MgCI 2 and K 2 S0 4 , as descπbed, for example, in G Kortum, Lehrbuch der Eiektrochemie [Textbook of Electrochemistry], 4th Edition, Verlag Chemie 1966, page 156 The measurement solution preferably comprises or consists of a body fluid It particularly preferably comprises or consists of blood The method can be carried out as an individual measurement or continuously The invention likewise relates to the use of an optical sensor descπbed above for optical pH determination of an aqueous measurement solution independently of the ion strength by the fluorescence method The examples below illustrate the invention Examples A 1-9. Preparation of the amtno-functional fluoresceins Example A1 Preparation of 12-azιdododecanoιc acid (1011 0 W / C-(CH 2 ) 11 -N 3 OH (10 1) 12-Bromododecanoιc acid (10 mmol) are dissolved in 30 ml of DMF, and 6 mol eq of sodium azide are added The suspension is stirred at 100°C for 16 hours, cooled and poured into 300 ml of ice water The aqueous phase is extracted three times with ether, and the organic phase is washed with saline solution, dried and evaporated The 2 2 g residue corresponds to 92 % of theory 1 H-NMR (CDCI 3 ) 3 26 (t, 2H, CH 2 N 3 ) IR (CDCI 3 ) 2098 cm' 1 (st , N 3 ) Example A2 Preparation of 12-azιdododecanoyl chloride (102) )MCH 2 ) ι r N cι 1 g ( 4 2 mmol) of 12-azιdododecanoιc acid is dissolved in 5 ml of methylene chloride, and 1 ml (12 mmol) of oxalyl chloride is added This solution is stirred for 4 hours, during which time a gas is evolved The solution is then evaporated, leaving a 1 3 g residue IR (CDCI 3 ) 2098 cm' 1 (st , N 3 ), 1796 cm" 1 (st , COCI) Example A3 Preparation of the compound of the formula (103) 2 43 g (7 mmol) of 4-amιnofluoresceιn (Fluka) and 6 ml of pyπdme are introduced into 300 ml of acetonitrile, and the mixture is cooled to 0°C The acid chloride from Example A2, dissolved in 40 ml of acetonitrile, is then added dropwise over the course of 30 minutes The mixture is then stirred at this temperature for 4 hours and filtered, and the filtrate is evaporated The residue is dissolved in 400 ml of methylene chloπde/methano! 80 20 and washed with 100 ml of 1 N HCI The organic phase is separated off and evaporated The crude product is purified by chromatography on silica gel using methylene chloπde/methanol 9 1 Yield 46 % m p 143°C FAB-MS 569 [M-H] " Example A4 Preparation of the compound of the formula (104) The compound from Example A3 is dissolved in MeOH, and 2 mol eq. of SnCI 2 H 2 0 are added, whereupon a gas is immediately evolved. The deep-red solution is stirred at RT for 5 hours, and aqueous ammonia is then added, whereupon a red solid precipitates. This is filtered off and dried in a high vacuum, m.p. > 270°C. Example A5 Preparation of the compound of the formula (105) .0, .OH N, ' O ' (105) 14.5 ml of triethylene glycol monochlorohydrin and 10 g of sodium azide are heated to 90°C without a solvent. After 24 hours, the mixture is cooled and filtered. The filtrate is distilled in a bulb tube at 0.1 mm and 120°C. Yield 67 %. FAB-MS: 176 [M+H] + . IR (KBr): 2109 (st., N 3 ). Example A6 Preparation of the compound of the formula (106) 0.188 mol of NaH are introduced into 150 ml of dry THF, and the product from Example A5 (0.171 mol) is added dropwise at 5°C. The mixture is then stirred at 0°C for 30 minutes. 38 ml (0.256 mol) of t-butyl bromoacetate in 60 ml of dry THF are then added dropwise. The reaction mixture is then stirred overnight and evaporated, and the residue is taken up in ether and washed with water The organic phase is washed with water and saline solution, dried and evaporated The residue is distilled at 0 09 mm and 140-150°C Yield 39 % 1 H- NMR (CDCI 3 ) 4 05 (s, 2H, CH 2 COOR), 1 50 (s, 9H, t-Bu) IR (KBr) 2106 (st , N 3 ), 1748 (st CO) Example A7 Preparation of the compound of the formula (107) O .0^ (107) OH 5 g (17 3 mmol) of the compound from Example A6 are mixed with 10 ml of water and 2 g of NaOH (50 mmol) The mixture is dissolved homogeneously by addition of ethanol and then warmed to 50°C After 30 minutes, the mixture is cooled and evaporated The residue is washed a number of times with ether, then dissolved in water and adjusted to pH 3 using 4n HCI The product is extracted with ether, and the organic phase is washed with saline solution, dried and evaporated, leaving 3 15 g (78 %) of a pale yellow oil Example A8 Preparation of the compound of the formula (108) 6 mmol of the compound from Example A7 are dissolved in 15 ml of dry THF, and 1 mol eq of carbonyldnmidazole is added After 4 hours at RT a solution of 4-amιnofluoresceιn in 100 ml of THF is added, and the mixture is stirred overnight, evaporated and purified on silica gel using methylene chloπde/methanol 95 5 to 85 15 Yield 64 % of orange crystals Example A9 Preparation of the compound of the formula (109) 0 177 mmol of the product from Example A8 is dissolved in 10 ml of methanol, and 0 48 mmol of SnCI 2 H 2 O is added, whereupon weak evolution of gas commences The mixture is subsequently adjusted to pH 9 using 1 N NaOH and then to pH 6 using 1 N HCI, during which the product precipitates The crystallization is completed by addition of THF and ether The solution is decanted off, and the residue is dried in a high vacuum FAB-MS 535 [M-H] " Examples B Preparation of thermoplastic, randomly segmented polvurethanes which are soluble in organic solvents Example B1 1 0 g of polytetrahydrofuran (PTHF), M n = 1000 g/mol), 1 0 g of polyethylene glycol (PEG, Mn = 1000 g/mol), 0 18 g of butanediol (BDO) and 0 005 mg of diazabicyclooctane (DABCO) are dissolved in 15 ml of dry THF under inert conditions (N 2 atmosphere) in a three-necked flask fitted with precipitation glass stirrer The mixture is allowed to react with 0 093 g of methylenediphenyl dnsocyanate (MDI) for 2 hours at 60°C 0 07 g of MDI are dissolved in 2 5 ml of THF in a 2nd three-necked flask (N 2 atmosphere), and a solution of 0 031 g of 4-amιnofluoresceιn in 5 ml of THF is added dropwise at RT The mixture is allowed to react for 60 minutes The yellow-fluorescent solution obtained is combmed with the first reaction solution, and the mixture is allowed to react at 60°C for 3 hours On pouring the polymer solution into 500 ml of methanol, the polymer is precipitated, and is then filtered off and dried at 20°C under reduced pressure The polymer is dissolved in 20 ml of THF and precipitated in 500 ml of methanol, this dissolution/precipitation operation is repeated, giving 2 24 g (71 % of theory) of yellow polymer Examples B2 to B7 are carried out analogously, again using the fluorescein dyes (104) and (109) from Examples A4 and A9 Polymers having the composition given in Table 1 are obtained Table 1 Polyurethanes containing covalently bound fluorescent dyes, amounts in g * PEO where M = 1000 1) 4'-Amιπofluoresceιn ** PEO where M = 2000 (104) 4'-(11-Amιnoιιndecylcarbonyl)- Tπol = 1,1 1-Trιs(hydroxymethyl)etrιane fluoresceinamine (109) 4'-(11-Amιno-2 5,8-trιoxo-decylcarbon- yl)fluoresceιnamιne Example C Production and charactenzation of the sensors Example C1 Glass substrates (plates with a diameter of 18 mm) are first cleaned in 30 % sodium hydroxide solution and then activated in 65 % nitric acid The activated plates are then silanized using 3-amιnopropyltπmethoxysιlane The polymer (5 %) from Example B5 is dissolved in tetrahydrofuran at from 20° to 25° and applied as a thin film to the silanized plate by spin coating at a speed of 500 revolutions per minute for 20 seconds The film is then dried for 12 hours at 60° under nitrogen The thickness of the membranes is about 1 μm Examples C2 and C3 The procedure is as in Example C1 using the corresponding polymers for Examples B6 and B7 Characterization of the sensor properties General procedure The sensors are installed in a flow cell The calibration or sample solutions are metered in by means of pumps and passed through the cell The measurement arrangement is thermostatted The light from a halogen lamp (white light, excitation wavelength 480 nm) is passed through an excitation filter, reflected at a dichroic mirror and bundled onto the planar sensors by means of lenses The fluorescent light (at 520 nm) emitted by the sensors is collected by the same lens system and passed through an emission filter and the dichroic mirror to a photodiode The fluorescence from the sensors is recorded during exposure by means of the calibration or sample solutions The pH can be determined directly from the measurement value Table 1 below shows the pH dependence of the ion strength of the electrolyte in the pH range from 6 7 to 8 0 obtained owing to the different membrane compositions Table 1 1 ) The numerical values relate to the difference in the pK a values measured once at an ion strength of 0 1 mol/l and once at an ion strength of 0 3 mol/l

Claims: WHAT IS CLAIMED IS. 1. An optical sensor for pH determination which is independent of the ion strength in the physiological range, comprising A) an transparent support material; B) a plasticizer-free, thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea which is soluble in organic solvents, formed from a) 10-40 % by weight of an aromatic, cycloaliphatic or linear aliphatic diisocyanate, b) 50-85 % by weight of a polyethylene glycol having a molecular weight of 2,000-10,000 daltons, c) 0-30 % by weight of a polytetrahydrofuran, aminopropyl-terminated polytetrahydrofuran, polypropylene glycol or ammopropyl-terminated polypropylene glycol having a molecular weight of 600-10,000 daltons, d) 0-10 % by weight of a linear or branched C 2 -C 12 alkylenedιol or C 2 -C 12 alkylenediamιne, e) 0-2 % by weight of a linear or branched C 3 -Cι 2 alkylenetrιol and f) 0 1-3 % by weight of a fluorescein dye containing an ammo or hydroxyl group bonded directly or at the end of a bridging group, where the percentages are based on the amount of polymer, and the amounts a) to f) total 100 2 An optical sensor according to claim 1 , wherein the amino or hydroxyl group of the fluorescein dye f) and the terminal isocyanate component a) form a urea or urethane group 3. An optical sensor according to claim 1 , wherein the diisocyanate, component a), in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is bis[4-ιsocyanatophenyl]methane (4,4'-MDI), 2,4- or 2,6-bιsιsocyanatotoluene (TDI), 1 ,6-bιsιsocyanatohexane (HDI), 5-isocyanato-3-isocyanatomethyl-1 ,1 ,3-trimethylcyclo- hexane (IPDI) or bιs[4-isocyanatocyclohexyl]methane (MDI), or a mixture of these diisocyanates. 4. An optical sensor according to claim 1 , wherein component b) in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is 70-85 % by weight of polyethylene glycol having a molecular weight of 600 - 2000 daltons. 5 An optical sensor according to claim 1 , wherein the diol, component d), in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is ethylene glycol, butanediol or hexanediol 6 An optical sensor according to claim 1 , wherein the alkylenetπol, component e), in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is 1 , 1 ,1 -Trιs(hydroxymethyl)ethane. 7 An optical sensor according to claim 1 , wherein component f) in the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea is 0 1 - 1 % of a fluorescein dye which is ammo- or hydroxyl-functionalized via a bridging group 8 An optical sensor according to claim 1 , wherein the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea has a molecular weight of from 10,000 to 30,000 daltons 9 An optical sensor according to claim 1 , wherein the thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea contains a fluorescein of the formula (la) or (lb) in which R^ is a bridging group and is linear or branched C 1 -C 20 alkylene, C 3 -C 20 alkenylene, C 3 -C 20 alkynylene or CH 2 -(O-(CHR 2 ) r -CH 2 -CH 2 ) n , R 2 is C r C 4 alkyl, X is NH 2 or OH, r is 0 or 1 , and n is a number from 1 to 20 10 An optical sensor according to claim 1 , wherein the transparent support material comprises glass, quartz or a thermoplastic or crosslmked plastic 1 1 An optical sensor according to claim 1 , wherein the thickness of the polymer layer B) is from 0 1 to 500 μm 12 An optical sensor according to claim 10, wherein the thickness of the polymer layer B) is from 1 to 100 μm 13 A method for the reversible, optical determination of the pH of an aqueous sample independently of ion strength by the fluorescence method, in which an optical sensor comprising A) a transparent support material, B) a plasticizer-free, thermoplastic, randomly segmented polyurethane, polyurea or polyurethane-urea which is soluble in organic solvents, formed from a) 10-40 % by weight of an aromatic, cycloaliphatic or linear aliphatic diisocyanate, b) 50-85 % by weight of a polyethylene glycol having a molecular weight of 2,000-10,000 daltons, c) 0-30 % by weight of a polytetrahydrofuran, ammopropyl-terminated polytetrahydrofuran, polypropylene glycol or ammopropyl-terminated polypropylene glycol having a molecular weight of 600-10,000 daltons, d) 0-10 % by weight of a linear or branched C 2 -C-| 2 alkylenedιol or C 2 -C 12 alkylenedιamιne, e) 0-2 % by weight of a linear or branched C 3 -C 12 alkylenetπol and f) 0 1-3 % by weight of a fluorescem dye containing an ammo or hydroxyl group bonded directly or at the end of a bridging group, where the percentages are based on the amount of polymer, and the amounts a) to f) total 100, is brought into contact with an aqueous measurement sample and exposed to excitation light, the fluorescence is measured, and the pH is calculated from the measured fluorescence intensity taking into account calibration curves 14 A method according to claim 13, wherein the measurement solution has a pH of from 6 7 to 7 8 15 A method according to claim 13, wherein the ion strength of the measurement solution is from 0 05 to 5 mol/l 16 A method according to claim 13, wherein the ion strength of the measurement solution is from 0 05 to 1 mol/l 17. A method according to claim 13, wherein the ion strength is built up essentially by 1 1 or 1.2 salts 18 A method according to claim 13, wherein the measurement solution comprises or consists of a body fluid. 19 A method according to claim 13, wherein the measurement solution comprises or consists of blood 20 The use of an optical sensor according to claim 1 for the optical determination of the pH of an aqueous measurement solution independently of the ion strength by the fluorescence method

Inventors: BARNARD STEVEN MARK, BECKELMANN DIRK, BERGER JOSEPH, ROUILLY MARIZEL, WALDNER ADRIAN, BARNARD, STEVEN, MARK, BECKELMANN, DIRK, BERGER, JOSEPH, ROUILLY, MARIZEL, WALDNER, ADRIAN
Applicants: BARNARD STEVEN MARK, BECKELMANN DIRK, BERGER JOSEPH, CIBA GEIGY AG, ROUILLY MARIZEL, WALDNER ADRIAN, BARNARD, STEVEN, MARK, BECKELMANN, DIRK, BERGER, JOSEPH, NOVARTIS AG, ROUILLY, MARIZEL, WALDNER, ADRIAN
Application year: 1997
Main classification: G01N31
IPC codes: G01J3, G01N21, G01N31, G01N33