Development of a biosensor specific for cysteine sulfoxides.
S-Alk(en)yl cysteine sulfoxides have been observed in several plants, mainly belonging to the onion family (Alliaceae), which are of high commercial interest (e.g. garlic, Allium sativum). The quality of most garlic containing herbal remedies produced from garlic powder is determined by their content of the cysteine sulfoxide alliin. Therefore, a comprehensive method for the documentation of alliin amounts present in the fresh plant material through to the final remedy is desirable. The newly developed biosensoric method described in this paper was designed in order to fulfil these demands. In contrast to conventional HPLC-methods, neither a pre-column derivatization nor a chromatographic separation are required allowing a high throughput of samples. This technique is based on immobilized alliinase (EC 4.4.1.4), which was combined with an ammonia-gas electrode. The enzyme was either placed in a small cartridge or was immobilized in direct contact of the electrode surface giving detection limits of 3.7 x 10(-7) and 5.9 x 10(-6) M. Founded on these experiments, a pH-sensitive electrolyte/insulator/semiconductor (EIS) layer structure made of Al/p-Si/SiO(2)/Si(3)N(4) was also combined with immobilized alliinase. Measurements could be performed in a range between 1 x 10(-5) and 1 x 10(-3) M alliin. All sensors were operated in the flow-through modus. A high specificity for alliin could be demonstrated for the electrode and a number of garlic samples were analyzed. Results gained with the new method showed a good correlation with those obtained with conventional HPLC-methods. In addition, onion and a variety of wild Allium species were analyzed in order to determine the amount of isoalliin or total cysteine sulfoxides present, respectively.
Keusgen
Michael
M
Institute for Pharmaceutical Biology, University of Bonn, Nussallee 6, D-53115, Bonn, Germany. m.keusgen@uni-bonn.de
Jünger
Martina
M
Krest
Ingo
I
Schöning
Michael J
MJ
D000490:Allium / Q000737:chemistry; D000641:Ammonia / Q000032:analysis; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D013437:Carbon-Sulfur Lyases / Q000737:chemistry*; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis / Q000737:chemistry; D003545:Cysteine / Q000031:analogs & derivatives* / Q000032:analysis*; D004800:Enzymes, Immobilized / Q000737:chemistry; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005737:Garlic / Q000737:chemistry; D028321:Plant Preparations / Q000032:analysis*; D010945:Plants, Edible / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013454:Sulfoxides / Q000032:analysis*
D020099:Coated Materials, Biocompatible; D004800:Enzymes, Immobilized; D028321:Plant Preparations; D013454:Sulfoxides; D000641:Ammonia; C006453:alliin; D013437:Carbon-Sulfur Lyases; C019243:alliin lyase; D003545:Cysteine