A New HPLC-ELSD Method To Quantify Indican in Polygonum tinctorium L. and To Evaluate β-Glucosidase Hydrolysis of Indican for Indigo Production

• Published in: Biotechnology progress Vol. 19; no. 6; pp. 1792 - 1797
• Main Authors: Angelini, Luciana G., Campeol, Elisabetta, Tozzi, Sabrina, Gilbert, Kerry G., Cooke, David T., John, Philip
• Format: Journal Article
• Language: English
• Published: USA American Chemical Society 01-11-2003; American Institute of Chemical Engineers
• Subjects: beta-Glucosidase - analysis; beta-Glucosidase - metabolism; Biological and medical sciences; Biotechnology; Chromatography, High Pressure Liquid - methods; Fundamental and applied biological sciences. Psychology; Hydrolysis; Indican - analysis; Indican - metabolism; Indigo Carmine; Indoles - metabolism; Photometry - methods; Plant Extracts - analysis; Plant Leaves - metabolism; Polygonum - metabolism; Polygonum tinctorium; Reproducibility of Results; Seasons; Sensitivity and Specificity; Indigoid dye; Enzyme; Polygonaceae; HPLC chromatography; Method; Hydrolysis; Polygonum; β-Glucosidase; Dicotyledones; Glycosidases; Angiospermae; Production; Hydrolases; Spermatophyta; O-Glycosidases; Quantitative analysis
• ISSN: 8756-7938; 1520-6033
• DOI: 10.1021/bp0300218

A method to quantify the indigo precursor indican (indoxyl‐β‐d‐glucoside) in Polygonum tinctoriumL. 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 P. tinctorium 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 P. tinctorium native β‐glucosidase to evaluate indigo production. Results showed that the ability to promote indigo formation increased as follows: almond β‐glucosidase ≤ Novarom G.