A RECONSIDERATION OF FRUCTAN BIOSYNTHESIS IN STORAGE ROOTS OF ASPARAGUS-OFFICINALIS L

Total soluble neutral carbohydrate (SNC) from storage roots of seven cultivars of Asparagus officinalis L. was analyzed by HPLC and TLC. Fructan accounted for roughly 90% of the total SNC and, of the fructan fraction, 81-98% was of degree of polymerization of five and larger (DP greater-than-or-equal-to 5). Smaller oligosaccharides and sucrose represented a correspondingly small proportion of the total SNC. This observation differs from the generally accepted view of Asparagus fructan, which emphasizes the presence of small oligofructan in root tissue. On the basis of chromatographic mobility, neokestose and isokestose were identified as the two components of the trisaccharide fraction. Kestose did not accumulate in any of the cultivars studied. A crude enzyme extract of Asparagus roots was prepared using established methods involving dialysis for 5 d. Microbiological analysis of this preparation revealed heavy bacterial contamination. Oligofructan apparently persisted in this protein preparation despite the long period of dialysis. These observations have consequences for the interpretation of previous reports of in vitro fructosyl transferase data for Asparagus root enzymes. The validity of long-term dialysis in the study of fructan enzymes is questioned. Crude protein extracts of roots were also desalted using a rapid gel-exclusion method. When incubated with sucrose under conditions similar to established techniques, both invertase and fructosyl transferase activities were detected in this preparation. The trisaccharide products of the fructosyl transferase reaction were identified on TLC as neokestose, isokestose and kestose; the last did not accumulate in vivo. Oligofructans of DP > 3 were also synthesized by this preparation and the pattern of products revealed by TLC was similar to previous reports. These oligofructan products of enzyme reactions were compared directly with the complete range of fructan structures accumulating in the tissue, using TLC. The enzyme products bore little resemblance to the native fructan pattern. Such enzyme activities are of doubtful physiological relevance and may be the result of artifactual or ancillary activities of other enzymes such as invertase. These findings are discussed with respect to previous studies of Asparagus root carbohydrates and serve as a basis for a critical reassessment of the enzymology of fructan biosynthesis both in this and other plant tissues.