Frozen aqueous systems

Paramagnetic species formed by reactions of the radiation-produced transient species in ice and frozen aqueous systems have been studied by ESR technique. The radiation-produced electrons have been found to react e.g. with acidic solutes to form H-atoms and with group 11(b) metal ions to give the corresponding univalent radical ions, while the holes can react with anions such as S04 2 and H2P04 giving the radical ions S04 and HP04. Evidence that the electron and hole are coupled to each other, and may in fact exist in irradiated pure ice primarily in an (exciton-like) bound state has been discussed. The present work provides evidence for the reactions of the radiation-produced positive holes apart from the reactions of the electrons. 

Figure 1. ESR spectra of y-irradiated frozen aqueous systems at 77°K. (a) H20 (b) 0.5M H2SOa ...

Table III. Comparison of the Relative Rates of Reactions of the Electron with Different Scavengers in Liquid and Frozen Aqueous Systems...

Figure 11. ESR spectra of y-irradiated frozen aqueous systems at 77°K. (a) 0.2M CiOa 2 (6) H2O (c) 0.1M CrO4-2, 2M NaOH after annealing (d) 2M NaOH after annealing (e) 0.1M CtOa 2, 2M HCIOa after annealing (f) 2M HCIOa after annealing.

The reverse action of a trypsin-free elastase isolated from porcine pancreas was studied in frozen aqueous systems and was found to catalyze peptide bond formation more effectively than in solution at room temperature (Haensler, 1998). The acceptance of free amino acids as nucleophilic amino components indicates a changed specificity of the endoprotease in frozen reaction mixtures. In elastase-catalyzed formation of Ser-, lie-, and Val-X-bonds in frozen aqueous reaction mix-... 

S. Gerisch, G. Ullmann, K. Stubenrauch, and H.-D. Jakubke, Enzymatic peptide synthesis in frozen aqueous systems influence of modified reaction conditions on the peptide yield, Biol. Chem. Hoppe Seyler, 1994, 375, 825-828. 

M. Wagner, H. J. Hofmann, and H.-D. Jakubke, Influence of freeze-concentration effect on proteinase-catalysed peptide synthesis in frozen aqueous systems, Biochim. Biophys. Acta 1997, 1338, 253-258. 

Schuster M, Aaviksaar A, Haga M, Ullmann U, Jakubke HD. Protease-catalyzed peptide synthesis in frozen aqueous systems the freeze concentration model. Biomed Biochim Acta 1991 50 S84-S89. 

Concomitant strand-break studies on frozen aqueous systems on irradiation suggest that both radical cations and radical anions can lead to strand breaks. They also show that DSBs are remarkably frequent under these conditions. This was interpreted in terms of proximal trapping of positive and negative radical centres, each leading to a strand break (Boon et al., 1984). 

A step-by-step peptide synthesis from the N- to the C-terminus is not possible with chemical methods as it risks partial epimerization due to the repeated carboxy activation procedures, In constrast, the stereo- and regiospecificity of serine and cysteine proteases ensures integrity of the stereogenic center and allows ecological reaction conditions without side-chain protection. Scheme 4 shows the synthesis scheme using clostripain and chymo-trypsin as catalysts.The second coupling reaction was carried out by enzyme catalysis in a frozen aqueous system (see Section 4.2.3.1). 

Enzymatic Peptide Synthesis in Frozen Aqueous Systems... 

Table 12.5-4. Comparative model peptide synthesis catalyzed by chymotrypsin in frozen aqueous systems and at room temperature.

Some new approaches to suppress competitive reactions in protease-catalyzed peptide synthesis have been developed in our group [14], namely leaving group manipulations at the acyl donor in kinetically controlled reactions, enzymatic synthesis in organic solvent-free microaqueous systems, cryoenzymatic peptide synthesis, and biotransformations in frozen aqueous systems using the reverse hydrolysis potential of proteases and other hydrolases... 

O. Fennema, Activity of enzymes in partially frozen aqueous systems. In Water Relations of Foods, R.B. Duckworth (ed.). Academic Press, London, 1975, pp. 397-413. 

Wojnarovits L, Takacs E (2008) Radiat Phys Chem 77 225 Wypych M (1999) Pulse radiolysis induced transients in frozen aqueous systems at low temperatures. In Mayer J (ed) Properties and reactions of radiation induced transients. Selected topics. Polish Scientific Publishers, Warszava, pp 5—37 Yang K, Manno PJ (1959) J Am Chem Soc 81 3507 Zador E, Warman JM, Hummel A (1973) Chem Phys Lett 23 363... 

Fennema, O. Actvity of enzymes in partially frozen aqueous systems. In Water relations of foods (Ed. Duckworth, R.B.), p. 397, Academic Press London-New York-San Francisco. 1975 Gachet, E., Martin, G.G., Vigneau, F., Meyer, G. Detection of genetically modified organisms (GMOs) by PGR a brief review of methodologies available. Trends Food Sci. Technol. 9, 380 (1999)... 

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