Abstract
A new colorimetric determination was devised for estimation of isothiocyanate content in radish root (Raphanus sativus L.) juice. Allylthiourea (AU) and trans-4-methylthio-3-butenyl thiourea (MBU) were prepared from allylisothiocyanate and Raphanus sativus L. cv. Aokubi Miyashige Daikon respectively.
It was observed that both thiourea derivatives turned blue with 25-fold diluted Grote reagent. In the absorption spectra of the colored solutions of AU and MBU, both peaks were observed at 605nm. After examination of several different conditions, new method based on the colorimetry of thiourea derivatives was established as follows. Each mixture of 1ml AU or 1ml MBU and 4ml modified Grote reagent was incubated at 37°C for 45min, and the optical density was determined at 600nm. The absorbancy was proportional to MBU concentration in the range between 20μg/ml and 200μg/ml.
After preliminary experiments, a suitable method was found to determine the isothiocyanate content in radish juice. Juice from radish root was kept at 30°C for 30min. To convert produced isothiocyanates into thiourea derivatives, a mixture of ethanol and aqueous ammonia was added to juice, followed by maintaining at 30°C for 60min. The mixture was neutralized with 50% acetic acid and filtered. An aliquot of the filtrate was employed for the determination of MBU as described previously. The resulting value was calculated as trans-4-methylthio-3-butenyl isothiocyanate.
Using the new method, comparison of the concentration of pungent principles in radish roots was performed, in respect to the differences in varieties and maturities as well as sections of the root.
