A New HPLC‐ELSD Method To Quantify Indican in <i>Polygonum tinctorium</i> L. and To Evaluate β‐Glucosidase Hydrolysis of Indican for Indigo Production

Abstract

<jats:title>Abstract</jats:title><jats:p>A method to quantify the indigo precursor indican (indoxyl‐β‐<jats:sc>d</jats:sc>‐glucoside) in <jats:italic>Polygonum tinctorium</jats:italic>L. has been developed. Plant material was extracted in deionized water, and indican was identified and quantified using high performance liquid chromatography (HPLC) coupled to an evaporative light scattering detector (ELSD). Results confirmed that with this method it is possible to measure indican content in a short time, obtaining reliable and reproducible data. Using this method, leaf indican content was quantified every 15 days during the growing season (from May to October) in <jats:italic>P. tinctorium</jats:italic> crops grown in a field experiment in Central Italy. Results showed that indican increased along the growing season until flowering and was positively affected by photosynthetic active radiation (PAR). Indican is naturally hydrolyzed by native β‐glucosidase to indoxyl and glucose, the indoxyl yielding indigo. The activity of two enzymes, sweet almond β‐glucosidase and Novarom G preparation, were compared with <jats:italic>P. tinctorium</jats:italic> native β‐glucosidase to evaluate indigo production. Results showed that the ability to promote indigo formation increased as follows: almond β‐glucosidase ≤ Novarom G.</jats:p>