Synthesis of 1-(4-Aminobutyl)-2-iminoimidazolidin-4-one Aimed at Preparation of a Creatinine-Specific Antibody

Abstract

For the purpose of obtaining a creatinine-specific antibody, a creatinine derivative with 4-aminobutyl, which was served as a linker for preparing the creatinine-bovine serum albumin (BSA) conjugate, was synthesized from 4-benzylaminobutan-1-ol in 8 steps. Production of anti-creatinine antibodies was observed in two rabbits using the creatinine-BSA conjugate, although their titer was rather low.Key words creatinine derivative; synthesis; guanidinylation; anti-creatinine immunoglobulin Recently, we developed an immunochromatographic system for determination of the N 1 ,N 12 -diacetyl spermine (DiAcSpm) concentration in human urine 1) (Fig. 1). DiAcSpm is a newly developed tumor marker and is unique among existing tumor markers in that it is sensitive in detecting early cancers. 2,3)The immunochromatographic testing device can be used for personal health control and would eventually contribute to improvement in the efficiency of early detection of cancer, which is important for reducing the number of fatal cases of cancer. The quantity of daily excretion of urine components is usually estimated from arbitrarily excreted spot urine samples by creatinine normalization and is expressed in nmol substance per g creatinine. The validity of creatinine normalization in estimating daily DiAcSpm excretion in urine was in fact confirmed, 4) and creatinine-normalized DiAcSpm in urine has been shown to be a promising tumor marker as described above. To determine the quantity of creatinine-normalized DiAcSpm by immunochromatography, it is necessary to simultaneously determine the quantities of creatinine in urine samples. Yaffe's reaction, which utilizes the reaction of creatinine with picric acid to form orange-red-colored creatinine picrate in an alkaline solution, can be used for this purpose.5) However, substances in urine such as pyruvate, glucose, ascorbate, and billirubin are also known to form colored complexes and interfere with accurate determination of creatinine. An enzymatic method utilizing creatinine imidohydrolase and glutamate dehydrogenase is more accurate 6) but is not readily applicable to creatinine assays on a solid support that are run in parallel with the immunochromatographic determination of DiAcSpm. A creatinine-specific antibody, if available, would be readily applicable to immunochromatographic determination of creatinine. Moreover, DiAcSpm and creatinine could be determined on the same strip by using gold nanoparticles labeled with two antibodies. To obtain a creatinine-specific antibody, creatinine as a hapten must be conjugated to an appropriate carrier macromolecule. With this in mind, we designed a creatinine derivative (10) with 4-aminobutyl.