Işınlama sonucunda Beta-Laktam grubu bazı antibiyotiklerde oluşan ara ürünlerin ESR yöntemi ile incelenmesi
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2005
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Hacettepe Üniversitesi, Fen Bilimleri Enstitüsü, Fizik Mühendisliği Anabilim Dalı
Abstract
İlaç üretiminde önde gelen önemli hususlardan biri de steril olmuş birim doz preparasyonlarının hazırlanmasıdır. Preparasyonların mikroorganizmalardan arındırılması klasik sterilizasyon teknikleri ile yapıldığında, özellikle sıcaklığa duyarlı ilaç ve ilaç etkin maddelerinde, ciddi boyutlarda degradasyonlar ortaya çıkabilmektedir. Bu sakıncayı ortadan kaldırmak ya da hiç değilse azaltmak için günümüzde yüksek enerjili radyasyonun kullanılması yoluna gidilmektedir. Başka deyimle günümüzde biyolojik kökenli ürünlerin ve ilaçların sterilizasyonu için daha çok yüksek enerjili radyasyonun kullanılması tercih edilmektedir. Bu tez çalışmasında, beta-laktam ailesine giren antibiyotiklerin hazırlanmasında ilaç etkin maddesi olarak kullanılan biri karbapenem (Meropenem trihidrat) diğer ikisi penisilin (piperasilin monohidrat ve sultamisilin tosilat) grubuna dahil olan üç bileşiğin radyasyon duyarlıkları incelenerek bu etkin maddeleri içeren ilaçların radyasyonla sterilizasyonlarının yapılıp yapılamayacakları araştırılmıştır. Bileşikler radyosterilizasyon için uluslararası kabul gören doz limitleri içerisinde kalınarak 1, 3, 6, 10 ve 15 kGy doz değerlerinde gamma ışınları ile ışınlanmış ve ışınlama sonucunda bu bileşiklerde oluşan radyolitik ara ürünlerin miktarları, mikrodalga doyum davranışları, sıcaklığın bu ara ürünler üzerine etkileri, odqa ve yüksek sıcaklıklardaki kararlılıkları ESR spektroskopisi yardımı ile incelenmiştir.
Işınlanan piperasilin monohidratın iki, sultamisilin tosilatın beş ve üçüncü bileşik olan meropenem trihidratın daha çok sayıda olmak üzere karmaşık sayılabilecek karakteristik rezonans tepesine sahip oldukları gözlenmiştir. Örneklerin ESR spektrumlarının mikrodalga gücüne bağlı değişimleri; 0,1-20 mW aralığında incelenerek spektruma katkı getiren olası radikal türleri belirlenmeye çalışılmıştır. Daha sonra sıcaklığın bu radikal türleri üzerine olan etkileri araştırılmış ve bu amaçla örnek sıcaklığı 100-410 K arasında değiştirilerek spektrumlarda oluşan olası değişimler gözlenmeye çalışılmıştır. Sinyal şiddetlerinin düşük sıcaklıklarda artarken yüksek sıcaklıklarda azaldıkları belirlenmiştir. Ancak, spektrumların genel görünüşlerinin değişmediği görülmüştür. Sinyal şiddetlerinde düşük sıcaklıklarda gözlenen artışların tersinir, yüksek sıcaklıklardaki azalmaların ise tersinir olmadığı saptanmıştır. İki ay süre ile oda sıcaklığında bekletilen örneklerin sinyal şiddetlerinde zamanla ortaya çıkan olası değişimler incelenmiş ve tüm rezonans tepeleri ile ilgili şiddetlerin başlangıçta hızlı bir sönüm gösterdikleri ancak zaman ilerledikçe bu sönüm hızının azaldığı gözlenmiştir. Sinyal şiddeti sönüm verilerine en iyi uyan kuramsal fonksiyonlar araştırılmış ve bu yolla spektruma katkı getiren radikal türleri ve bunların oda sıcaklığındaki sönüm sabitleri belirlenmiştir.
Yüksek sıcaklıklarda değişik sürelerle tavlanan örneklerin sinyal şiddetlerinde gözlenen azalmalar dikkate alınarak deneysel spektrumun oluşumuna katkı getiren radikal türlerinin sönüm aktivasyon enerjileri hesaplanmış ve bulunan değerlerin benzer molekül yapısına sahip olan bileşikler için literatürde elde edilen değerlerle uyumlu olduğu belirlenmiştir. Ancak, sultamisilin tosilat örneği ile ilgili tavlama bulgularına anlamlı sayılabilecek bir veri uyarlama işlemi yapılamadığından, bu bileşikte oluşan kökçe türlerinin aktivasyon enerjileri hesaplanamamıştır. Ayrıca tüm örnek türleri için doz-cevap eğrileri oluşturulmuş, bu eğrilerden yararlanarak incelenen bileşiklerin dozimetre malzemesi olarak kullanılıp kullanılamayacakları araştırılmıştır. Meropenem trihidrat bileşiğinin bu amaçla kullanılabileceği sonucuna varılmıştır.
Tüm bu incelemelerin sonucunda, meropenem trihidrat örneği ESR spektrumunun oluşumuna değişik ağırlık oranlarında olmak üzere dört; piperasilin monohidrat ve sultamisilin tosilatın spektrumlarına da yine değişik ağırlık oranlarında olmak üzere iki farklı türde radikalin katkı getirdikleri yargısına varılmıştır. Moleküler yapılar dikkate alınarak bu radikallerin oluşum tepkimeleri için öneriler yapılmıştır. Önerilen radikal türlerinin ve bunların oluşum mekanizmalarının ancak spektrum simülasyon hesaplamalarının tamamlanması ile kesinlik kazanabileceğini belirtmek gerekir. Oldukça düşük sayılabilecek radyasyon verimine sahip olmaları nedeniyle her üç etkin maddeyi içeren ilaçların radyasyonla sterilizasyona uygun oldukları sonucuna varılmıştır.
Particular interest now centers on the preparation of steril-unit dose preparations. When preparations are purified from microorganisms using classic sterilization techniques, serious degradations may occur especially in temperature sensitive drugs and drug active components. In order to remove or at least reduce this drawbacks, use of high energy radiation now become popular. In other words, the use of high energy radiation for sterilization of bioproducts and pharmaceuticals is now preferred to the other techniques. In this M.S. work, radiation sensitiveness of three compounds, one of them is included in carbapenems and the others are included in penicillins groups which used as active drug components while preparing beta-lactam antibiotics were investigated and searched whether these drugs can be sterilized by radiation or not. Compounds were irradiated by gamma rays at 1,3,6,10 and 15 kGy irradiation doses. These dose values stay in the international dose limits accepted for radiation sterilization. After that, the amounts, microwave saturation behaviours, stability at room and high temperatures of the radiolytic intermediates produced in the samples after irradiation and effects of temperatures on these intermediates were investigated by Electron Spin Resonance (ESR) spectroscopy. It is observed that the irradiated piperacillin monohydrate has two, sultamicillin tosylate has five and the third sample called meropenem trihydrate has still much more characteristic resonance peaks making complex the ESR spectra. The microwave dependent variations of ESR spectra for all the samples were examined in the range of 0.1-20 mW to determine the possible radical species contributing to the experimental spectra. Afterwads, the effects of temperature on these radical species were investigated and for this aim, possible changes arising in spectra were tried to observe while changing sample temperature in the range of 100 and 410 K. While the signal intensities of the spectra were increased at low temperatures, they were found to decrease at high temperatures. However, it was observed that the general patterns of the spectra did not change. Whereas the increases of the signal intensities at low temperatures were reversible, their decreases at high temperatures were not. The samples were stored at room temperature for two months and the changes in the signal were examined. The intensities related to all resonance peaks showed fast decay at the beginning of storage period, but as time passed the rate of these decays were observed to decrease. Theoritical functions describing well the signal intensity decay data were searched and by this way radical species contributing to experimental spectra and their room temperature decay constants were determined. The decay activation energies of the radical species contributing to ESR spectra were calculated by taking into consideration the signal intensities decay data of the studied compounds annealed at high temperatures for different period of time and it was found that the calculated values were correlated well with those obtained in the literature for samples exhibiting similar molecular structure. However, it was not possible to calculate activation energies of radical species produced in sultamicillin tosylate due to lack of reliable signal intensity decay data at high temperature for this compound. Moreover, the dose-response curves were constructed for all the sample species and by using the data obtained from these curves, an evaluation was made whether investigated compounds can be used or not for dosimetric purposes. It was concluded that meropenem trihydrate can be used for this purpose. As a result of these investigations, it was concluded that, four and two different radical species contributed with different weights to the formation of experimental ESR spectra of meropenem and both piperacillin monohydrate and sultamicillin tosylate, respectively. By taking into consideration molecular structures, different propositions were made concerning the formation reactions of these radicals. It is needed to state that proposed radical species and their formation mechanisms can only become definite after completion of spectrum simulation calculations. However, it was found that the drugs including these compounds as active ingredients were suitable for radiation sterilization since they have rather low radiation yield.
Particular interest now centers on the preparation of steril-unit dose preparations. When preparations are purified from microorganisms using classic sterilization techniques, serious degradations may occur especially in temperature sensitive drugs and drug active components. In order to remove or at least reduce this drawbacks, use of high energy radiation now become popular. In other words, the use of high energy radiation for sterilization of bioproducts and pharmaceuticals is now preferred to the other techniques. In this M.S. work, radiation sensitiveness of three compounds, one of them is included in carbapenems and the others are included in penicillins groups which used as active drug components while preparing beta-lactam antibiotics were investigated and searched whether these drugs can be sterilized by radiation or not. Compounds were irradiated by gamma rays at 1,3,6,10 and 15 kGy irradiation doses. These dose values stay in the international dose limits accepted for radiation sterilization. After that, the amounts, microwave saturation behaviours, stability at room and high temperatures of the radiolytic intermediates produced in the samples after irradiation and effects of temperatures on these intermediates were investigated by Electron Spin Resonance (ESR) spectroscopy. It is observed that the irradiated piperacillin monohydrate has two, sultamicillin tosylate has five and the third sample called meropenem trihydrate has still much more characteristic resonance peaks making complex the ESR spectra. The microwave dependent variations of ESR spectra for all the samples were examined in the range of 0.1-20 mW to determine the possible radical species contributing to the experimental spectra. Afterwads, the effects of temperature on these radical species were investigated and for this aim, possible changes arising in spectra were tried to observe while changing sample temperature in the range of 100 and 410 K. While the signal intensities of the spectra were increased at low temperatures, they were found to decrease at high temperatures. However, it was observed that the general patterns of the spectra did not change. Whereas the increases of the signal intensities at low temperatures were reversible, their decreases at high temperatures were not. The samples were stored at room temperature for two months and the changes in the signal were examined. The intensities related to all resonance peaks showed fast decay at the beginning of storage period, but as time passed the rate of these decays were observed to decrease. Theoritical functions describing well the signal intensity decay data were searched and by this way radical species contributing to experimental spectra and their room temperature decay constants were determined. The decay activation energies of the radical species contributing to ESR spectra were calculated by taking into consideration the signal intensities decay data of the studied compounds annealed at high temperatures for different period of time and it was found that the calculated values were correlated well with those obtained in the literature for samples exhibiting similar molecular structure. However, it was not possible to calculate activation energies of radical species produced in sultamicillin tosylate due to lack of reliable signal intensity decay data at high temperature for this compound. Moreover, the dose-response curves were constructed for all the sample species and by using the data obtained from these curves, an evaluation was made whether investigated compounds can be used or not for dosimetric purposes. It was concluded that meropenem trihydrate can be used for this purpose. As a result of these investigations, it was concluded that, four and two different radical species contributed with different weights to the formation of experimental ESR spectra of meropenem and both piperacillin monohydrate and sultamicillin tosylate, respectively. By taking into consideration molecular structures, different propositions were made concerning the formation reactions of these radicals. It is needed to state that proposed radical species and their formation mechanisms can only become definite after completion of spectrum simulation calculations. However, it was found that the drugs including these compounds as active ingredients were suitable for radiation sterilization since they have rather low radiation yield.
Description
Keywords
Electron spin resonance (ESR), Elektron spin rezonansı (ESR), Sterilization, Sterilizasyon, Radiation sterilization, Radyasyon sterilizasyonu, Radyasyonla sterilizasyon, Beta-Lactam antibiotics, Beta-Laktam antibiyotikler, Meropenem trihydrate, Meropenem trihidrat, Piperacillin monohydrate, Piperasilin monohidrat, Sultamicillin tosylate, Sultamisilin tosilat
Citation
Tepe, S. (2005). Işınlama sonucunda Beta-Laktam grubu bazı antibiyotiklerde oluşan ara ürünlerin ESR yöntemi ile incelenmesi. (Yayımlanmamış yüksek lisans tezi). Ankara : Hacettepe Üniversitesi, Fen Bilimleri Enstitüsü, Fizik Mühendisliği Anabilim Dalı.