Plant cell culture for production of natural products with reduced flucosinolate contamination

Abstract

Disclosed are methods for obtaining a natural product preparation with reduced glucosinolate contamination from a plant of Brassicaceae. The methods can include cultivating a plant callus from a plant capable of producing a desired natural product, selecting a callus with reduced glucosinolate production, and cultivating the selected callus in a liquid medium. The method can also include recovering the natural product from the culture. Also disclosed are methods for obtaining cabbage anthocyanin with reduced glucosinolate contamination. The methods can include cultivating a red cabbage plant callus with reduced glucosinolate production in a liquid medium to obtain a suspension culture and cultivating the suspension culture in a medium lacking a nitrogen source. The method can also include recovering the anthocyanin with reduced glucosinolate contamination from the culture. Finally, several specific low-glucosinolate cell lines are described.

What is claimed is: 
     
         1 . An isolated cabbage cell line, comprising callus cells derived from a  Brassica  plant, the callus cells being selected to grow in suspension cell culture and to produce less than 76.54 μg total glucosinolate per gram fresh cell weight and at least 81.9 mg of anthocyanin per liter of packed cells when grown in cell suspension culture. 
     
     
         2 . The isolated cabbage cell line of  claim 1 , wherein the callus cells are selected to produce anthocyanins at a ratio of anthocyanin to glucosinolate of at least 40:1. 
     
     
         3 . The isolated cabbage cell line of  claim 1 , wherein the callus cells are selected to have reduced glucosinolate production of at least a 10% decrease as compared to the  Brassica  plant from which the callus cells were derived. 
     
     
         4 . The isolated cabbage cell line of  claim 1 , wherein the callus cells are selected to have reduced glucosinolate production of less than 5 μg of total glucosinolate per gram fresh cell weight. 
     
     
         5 . The isolated cabbage cell line of  claim 1 , wherein the callus cells are selected to produce at least 1119.0 mg of anthocyanin per liter of packed cells when grown in cell suspension culture. 
     
     
         6 . A cell suspension culture, comprising:
 the callus cells of  claim 1 ; and   a liquid suspension medium,   the callus cells growing in the liquid suspension medium and producing less than 76.54 μg total glucosinolate per gram fresh cell weight and at least 81.9 mg of anthocyanin per liter of packed cells.   
     
     
         7 . An isolated cabbage cell line, comprising callus cells derived from a  Brassica  plant, the callus cells being selected to produce less than 76.54 μg total glucosinolate per gram fresh cell weight and anthocyanins at a ratio of anthocyanin to glucosinolate of at least 40:1. 
     
     
         8 . The isolated cabbage cell line of  claim 7 , wherein the callus cells are selected to produce anthocyanins at a ratio of anthocyanin to glucosinolate of at least 100:1. 
     
     
         9 . The isolated cabbage cell line of  claim 7 , wherein the callus cells are selected to produce at least 81.9 mg of anthocyanin per liter of packed cells when grown in cell suspension culture. 
     
     
         10 . The isolated cabbage cell line of  claim 7 , wherein the callus cells are selected to have reduced glucosinolate production of at least a 10% decrease as compared to the  Brassica  plant from which the callus cells were derived. 
     
     
         11 . A cell suspension culture, comprising:
 the callus cells of  claim 7 ; and   a liquid suspension medium,   the callus cells growing in the liquid suspension medium and producing less than 76.54 g total glucosinolate per gram fresh cell weight and anthocyanins at a ratio of anthocyanin to glucosinolate of at least 40:1.   
     
     
         12 . An isolated cabbage cell line, comprising callus cells derived from a  Brassica  plant, the callus cells being selected to have reduced glucosinolate production of at least a 10% decrease as compared to the  Brassica  plant from which the callus cells were derived and to produce anthocyanins at a ratio of anthocyanin to glucosinolate of at least 40:1. 
     
     
         13 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to produce anthocyanins at a ratio of anthocyanin to glucosinolate of at least 100:1. 
     
     
         14 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to have reduced glucosinolate production of less than 76.54 μg of total glucosinolate per gram fresh cell weight. 
     
     
         15 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to have reduced glucosinolate production of less than 5 μg of total glucosinolate per gram fresh cell weight. 
     
     
         16 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to produce at least 81.9 mg of anthocyanin per liter of packed cells when grown in cell suspension culture. 
     
     
         17 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to produce at least 1119.0 mg of anthocyanin per liter of packed cells when grown in cell suspension culture. 
     
     
         18 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to have reduced glucosinolate production of at least a 50% decrease as compared to the  Brassica  plant from which the callus cells were derived. 
     
     
         19 . The isolated cabbage cell line of  claim 12 , wherein the callus cells are selected to have reduced glucosinolate production of at least a 90% decrease as compared to the  Brassica  plant from which the callus cells were derived. 
     
     
         20 . A cell suspension culture, comprising:
 the callus cells of  claim 12 ; and   a liquid suspension medium,   the callus cells growing in the liquid suspension medium and having reduced glucosinolate production of at least a 10% decrease as compared to the  Brassica  plant from which the callus cells were derived and producing anthocyanins at a ratio of anthocyanin to glucosinolate of at least 40:1.