Potenciometrično določanje in študij interakcij kalcijevih ionov s trinatrijevim citratom in humanim albuminom

Kastelec, Lea (author), Kolar, Mitja (mentor), Novljan, Gregor (co-mentor)

Določanje kalcijevih ionov v bioloških matricah (serum, kri, urin itd.) je možno z uporabo klasičnih in instrumentalnih metod. Pri "in vitro" študijah interakcij kalcija s trinatrijevim citratom in humanim albuminom je najprimernejša metoda potenciometrija z uporabo kalcijeve iono-selektivne elektrode (Ca/ISE). Ca/ISE predstavlja najboljšo kombinacijo odzivnosti, občutljivosti, specifičnosti in ponovljivosti za "in vitro" model kvantitativnega spremljanja sprememb koncentracij kalcijevih ionov ob konici dializnega katetra v resničnem času. V serumu je kalcij ioniziran, kompleksiran ali vezan na različne proteine, med katerimi prevladuje albumin. Nemoteno delovanje človeškega telesa je pogojeno z vzdrževanjem normalnega razmerja serumske koncentracije kalcija. Zaradi izhajanja trinatrijevega citrata iz konice katetra lahko pride do znižanja koncentracij ioniziranega kalcija v telesu. V sklopu magistrskega dela smo izvajali eksperimente pri različnih pogojih npr.: v odprtem sistemu pri sobni temperaturi, v zaprtem sistemu pri 36,5 °C, v ali ob odsotnosti zraka, ob prisotnosti albumina itd. Z izbiro ustrezne Ca/ISE in kalibracijo v izbranem koncentracijskem območju smo zagotovili točnost, natančnost in ponovljivost meritev. Pri poskusih smo zaradi dobre ponovljivosti in majhnega odstopanja izmerjenega naklona od teoretične vrednosti uporabili Mettler Toledo perfectIONTM Ca/ISE. Odzivni čas elektrode je 100 s, njen naklon v zaprtem sistemu je 29,3 ± 0,3 mV, kar je zelo blizu Nernstovega naklona. Potrdili smo vpliv temperature na odziv Ca/ISE; z višanjem temperature narašča tudi naklon elektrode. Poleg sprememb koncentracij kalcijevih ionov ob izhajanju trinatrijevega citrata iz konice katetra smo merili tudi pH. Potrdili smo mehanizem vezave kalcija na albumin ter ovrednotili vpliv kislinsko-bazičnega ravnotežja – pri višjem pH se je koncentracija prostih kalcijevih ionov znižala zaradi njihove vezave na humani albumin.

• Language: Slovenian
• Keywords: iono-selektivna elektroda, kalcij, natrijev citrat, humani albumin, hemodializa
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: FKKT - Faculty of Chemistry and Chemical Technology
• Year: 2019
• PID: 20.500.12556/RUL-113241
• COBISS.SI-ID: 1538506435
• Publication date in RUL: 16.12.2019

Secondary language

• Language: English
• Title: Potentiometric determination and study of interaction of calcium ions with trisodium citrate and human albumin

Abstract

Determination of calcium ions in biological matrices (serum, blood, urine etc.) is possible with the use of classical and instrumental methods. With in vitro studies of interactions of calcium with trisodium citrate and human albumin, the preferred method is potentiometry with the use of calcium ion-selective electrode (Ca/ISE). Ca/ISE presents the best combination of responsiveness, sensitivity, specificity and repeatability for the in vitro model of quantitative monitoring of changes of calcium ions concentrations at the tip of the dialysis catheter in real time. In the serum, calcium is ionised, complexed or bound to different proteins, among which albumin predominates. Smooth functioning of the human body is contingent on maintaining a normal ratio of serum concentration of calcium. The leakage of trisodium citrate from the tip of the catheter can lead to reduced concentrations of ionised calcium in the body. Within this master’s thesis, experiments have been carried out under different conditions, for instance in an open system at room temperature, in a closed system at 36.5 °C, in the absence of air, in the presence of albumin etc. With the choice of an appropriate Ca/ISE and calibration in the chosen concentration range, accuracy, precision and repeatability of the measurements have been ensured. In experiments, Mettler Toledo perfectIONTM Ca/ISE has been used due to good repeatability and small deviation of the measured slope from the theoretical value. The response time of the electrode is 100 s, its slope in the closed system is 29,3 ± 0,3 mV, which is very close to the Nernst slope. The influence of temperature on the reaction of Ca/ISE has been confirmed; increase in temperature results in the increase of the electrode slope. In addition to the changes of calcium ions concentrations after the leakage of trisodium citrate from the tip of the catheter, also pH has been measured. The mechanism of calcium binding to albumin has been confirmed and the influence of the acid-base balance evaluated – with a higher pH, the concentration of free calcium ions decreased due to their binding to human albumin.

• Keywords: ion-selective electrode, calcium, sodium citrate, human albumin, haemodialysis