Immobilization of Low-Cost Alternative Vegetable Peroxidase (Raphanus sativus L. peroxidase): Choice of Support/Technique and Characterization.

In the present work the radish (Raphanus sativus L.) was used as the low-cost alternative source of peroxidase. The enzyme was immobilized in different supports: coconut fiber (CF), calcium alginate microspheres (CAMs) and silica SBA-15/albumin hybrid (HB). Physical adsorption (PA) and covalent binding (CB) as immobilization techniques were evaluated. Immobilized biocatalysts (IBs) obtained were physicochemical and morphologically characterized by SEM, FTIR and TGA. Also, optimum pH/temperature and operational stability were determined. For all supports, the immobilization by covalent binding provided the higher immobilization efficiencies-immobilization yield (IY%) of 89.99 ± 0.38% and 77.74 ± 0.42% for HB and CF, respectively. For CAMs the activity recovery (AR) was of 11.83 ± 0.68%. All IBs showed optimum pH at 6.0. Regarding optimum temperature of the biocatalysts, HB-CB and CAM-CB maintained the original optimum temperature of the free enzyme (40 °C). HB-CB showed higher operational stability, maintaining around 65% of the initial activity after four consecutive cycles. SEM, FTIR and TGA results suggest the enzyme presence on the IBs. Radish peroxidase immobilized on HB support by covalent binding is promising in future biotechnological applications.

(99m)Tc complexes with activated ester functions; ligands comprising a 3,4-diamino-benzoate backbone.

Preformed (99m)Tc chelates with an activated ester function are useful for the gentle labeling of proteins (precomplexation route). In this context, new heterobifunctional ligands derived from 2,3,5,6-tetrafluorophenyl (TFP) 3,4-diamino-benzoates (OC1, OC3, OC4) were synthesized. Their corresponding (99m)Tc-complexation and protein-conjugation characteristics were elucidated and compared with the results previously reported using 2,3,5,6-tetrafluorophenyl N-(S-benzoylthioacetyl)glycylglycyl-p-aminobenzoate (OC2). The reaction temperatures and the reaction time markedly influenced complexation yields. Compared with OC2, the (99m)Tc-chelate formation with OC1 and OC4 was more effective, showing radiochemical yields of 60% and 70% within 20 min, respectively. Owing to steric hindrance, the complexation of OC3, however, did not exceed 10%. No-carrier-added (99m)Tc complexes were conjugated at pH 10 with the anti-EGF-receptor monoclonal antibody MAb425, resulting in labeling yields of 14% for (99m)Tc-OC1 and 7% for (99m)Tc-OC4 after incubating for 20 min at 30 degrees C. Increasing the temperature to 40 degrees C improved these results by 14% and 3%, respectively. As compared with (99m)Tc-OC2, which provides the chelating substituent at the 4-phenyl position only, the application of 3,4-phenyl substituents proved less appropriate for protein conjugation. However, the 3,4-diaminobenzoate backbone may be attractive for an alternative design of novel (99m)Tc N2S2 or N3S complexes, because they show excellent complexation characteristics.