Hawthorn extract has a significant amount of flavonoid and is of high importance due to the antioxidant property. Bioassay is a biological testing procedure for estimating the concentration of a substance. There are various methods for measuring antioxidant biological compounds. Qualitative and quantitative techniques of polyphenols measurement include chromatography, HPLC and GC-MS. These techniques are expensive and time-consuming so the development of biosensors can overcome these limitations. In order to prevent damage by free radicals, the body has a defense system of antioxidants. Plant bionanosensor is a certain type and novel approach of biosensors that has been fabricated by this author for the first time. The study was used to determine two types of flavonoids concentrations in vitro (35%ethanol 65% water) in hawthorn extract through rotation with three replications by sas9.1 software. The different levels of flavonoid rotation are significant with a probability of 99%. So far, no report was made about such a plant bionanosensor at international level from other researchers