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Merjenje pO2 v ustni sluznici podgan po nanosu hiperemikov z EPR oksimetrijo
Avtor: Bizimoska Saška
Mentor: doc. dr. Milan Petelin, dr. stom.
IZHODIŠCE. Mazilne podlage, ki se uporabljajo kot nosilci ucinkovin za zdravljenje ustnih bolezni, morajo imeti sposobnost dobre adhezije na ustno sluznico, sicer jih odplavi tok sline. Poznanih je vec mazilnih podlag, kot npr. Orabaza, Carbopol in polimetil-metakrilat. V dosedanjih raziskavah omenjenih podlag, ki so bile narejene v in vitro pogojih, je najboljše bioadhezivne lastnosti pokazal polimetil-metakrilat.
NAMEN. Namenili smo se primerjati nekatere lastnosti treh mazilnih podlag (Orabaze, Carbopola, polimetil-metakrilata) v in vivo pogojih. Z dodajanjem hiperemika v izbrane podlage smo v ustni sluznici povzrocili spremembo pO2, ki smo jo merili z metodo EPR.
HIPOTEZA. Postavili smo si naslednji delovni hipotezi:
- hipoteza 1: polimetil-metakrilat bo tudi v in vivo pogojih podlaga, ki bo omogocila najvecjo in najdlje casa trajajoco spremembo pO2 v ustni sluznici;
- hipoteza 2: s spreminjanjem koncentracije hiperemika v mazilni podlagi lahko dosešemo razlicne nivoje pO2 v ustni sluznici.
METODE. Raziskavo smo izvedli na samicah podgan soja Wistar. Primerjali smo tri podlage z bioadhezivnimi lastnostmi (Orabaza, Carbopol, polimetil-metakrilat) in podlago brez bioadhezivnih lastnosti (Miglyol). Vsaki smo dodali hiperemik (benzil-nikotinat), da je nastala 2% mazilna raztopina. Slednjo smo nato v kolicini 0,05 ml nanesli na licno sluznico podgan, v katero smo predhodno pod epitelij vstavili spinski oznacevalec. Kontrolnim skupinam smo nanašali samo podlage brez hiperemika. Po nanosu smo z EPR spektrometrom, prirejenim za delovanje pri 1,2 GHz, spremljali casovni potek sprememb pO2 v ustni sluznici podgan. Podlago, ki je omogocila najvecji ucinek hiperemika (polimetil-metakrilat), smo nadalje izbrali za testiranje druge hipoteze. Lego spinskega oznacevalca in spremembe ob njem smo ocenili tudi histološko. Rezultate smo statisticno obdelali z analizo variance in post hoc Tukey-evim testom.
REZULTATI. Ucinek anestezije na izmerjeni pO2 je najmanjši v obmocju telesne temperature živali med 36,6 °C in 37,2 °C (k=1,09; r=0,96). Izbrane podlage so se medsebojno razlikovale glede casa, ko je bila dosežena najvecja sprememba pO2 in glede površin pod krivuljami. Primerjava casov ob najvecji spremembi pO2 je pokazala statisticno znacilne razlike med posameznimi podlagami (p<<0,01; post hoc Tukey-ev test), razen med Orabazo in Miglyolom. Glede površin pod krivuljami se je statisticno znacilno od ostalih podlag razlikoval le polimetil-metakrilat (p<<0,01; post hoc Tukey-ev test). Po nanosu razlicnih koncentracij hiperemika v polimetil-metakrilatu so se skupine živali medsebojno razlikovale glede najvecje povzrocene spremembe pO2 v ustni sluznici. Statisticno znacilne razlike smo ugotovili med 2% in 3%, 2% in 3,5% ter 2% in 4% koncentracijo hiperemika (p=0,03; post hoc Tukey-ev test).
ZAKLJUCKI. V raziskavi smo prvi prikazali uporabnost metode EPR oksimetrije za merjenje pO2 v ustni sluznici podgan v in vivo pogojih. Metoda omogoca, ob upoštevanju dolocenih omejitev, preucevanje casovnega poteka delovanja ucinkovin, vgrajenih v razlicne mazilne podlage. Rezultati so potrdili naši hipotezi. S primerjavo izbranih mazilnih podlag z dodanim hiperemikom smo ugotovili, da je najvecjo in najdlje casa trajajoco spremembo pO2 povzrocil hiperemik v polimetil-metakrilatu. Z zmernim povišanjem koncentracije hiperemika v polimetil-metakrilatu sicer lahko dosežemo višji pO2 v ustni sluznici živali, vendar le do praga nasicenja.
[Abstract / English version]
Measuring of pO2 in mounth mucosa of rats after adding the hyperemics with EPR oxyomerty
Author: Bizimoska Saška
Mentor: doc. dr. Milan Petelin, dr. stom.
BACKGROUND. Ointments acting as vehicles for local delivery of drugs to the oral mucosa need to have accellent adhesive properties due to the flow of saliva. Such ointments are Orabase, Carbopol and polymethyl methacrylate. Many different researches on previously mentioned ointments have been made so far, mostly in vitro, where polymethyl methacrylate has showed the best results on bioadhesion.
AIM. Our research set to compare three adhesive ointments (Orabase, Carbopol, polymethyl methacrylate) in vivo. We added a hyperemic to each of the chosen ointments, thereby we caused a change in local pO2 in oral mucosa, which we then followed by EPR.
HYPOTHESIS. We tested these two hypotheses:
- hypothesis 1: polymethyl methacrylate will make, in vivo as well, the greatest and longest change in pO2 in oral mucosa possible,
- hypothesis 2: different levels of pO2 in oral mucosa can be reached by changing the concentration of the hyperemic.
METHODS. Our research was conducted on female Wistar rats. We compared three ointments with bioadhesive properties (Orabase, Carbopol, polymethyl methacrylate) and the ointment without such properties (Miglyol). We added the hyperemic (benzil-nicotinate) to each of them and then applied 0,05 ml of the 2% solution to rats' buccal mucosa, where we had previously put a paramagnetic probe. Rats in control groups were given only ointments without the hyperemic. After the application we followed the time-course of the pO2 changes in oral mucosa with the EPR spectrometer, adjusted for operating at 1,2 GHz. The ointment which made possible for the hyperemic to cause the greatest change in pO2 (polymethyl methacrylate) was chosen to test the second hypothesis. The position of the paramagnetic probe and changes in its surroundings were also assessed histologically. Results were analyzed using the analysis of variance followed by post hoc Tukey's test.
RESULTS. The effect of anesthesia on measured pO2 is smallest in the body temperature range of 36,6 °C to 37,2 °C (k=1,09; r=0,96). There were differences among chosen ointments in the time, when the greatest change in pO2 was established, and in the areas under the curves. Comparing the time at the greatest change of pO2, we found statistically significant differences between individual ointments (p<<0,01; post hoc Tukey's test) except among Orabase and Miglyol. Considering the areas under the curves, there were statistical differences only when comparing each ointment to polymethyl methacrylate (p<<0,01; post hoc Tukey's test). After applying different concentrations of the hyperemic in polymethyl methacrylate, we found that individual groups differed in the greatest change of pO2 established in oral mucosa. There were statistically significant differences between 2% and 3%, 2% and 3,5% and 2% and 4% concentration of the hyperemic (p=0,03; post hoc Tukey's test).
CONCLUSIONS. We were the first to show in our research the usefulness of EPR oximetry for measuring pO2 in rats' oral mucosa in vivo. This method is suitable for following the time-course of the effect of drugs in different adhesive ointments. The results confirmed our hypotheses. The greatest and longest change in pO2 was caused by the hyperemic in polymethyl methacrylate. Higher pO2 in oral mucosa can be established by increasing the concentration of the hyperemic in polymethyl methacrylate, but only until saturation is reached.