Summary• An analysis of fructan structures, to increase the understanding of biosynthetic pathways and enzymology of fructan synthesis in root and leaf tissues of Lolium perenne is reported.• Fructan extracted from stubble of L. perenne plants was analyzed by high performance anion exchange chromatography and pulsed amperometric detection (HPAEC-PAD) using a new desalting technique. Structures of fructan isomers, separated up to DP16 (DP, degree of polymerization), were established by chromatographic elution times or by GC-MS.• Fructans of DP8 belonged essentially to three series: inulin series, inulin neoseries and the levan neoseries, which is/are different in glucose (terminal or internal) and linked fructose residues. High DP fructans (DP > 8) comprised 75% molecules with an internal glucose residue. They had some branch points although 1 and 6 kestotetraose could not be detected and the β (2 -6) linked fructose residues were 70 times more abundant than the β (2 -1) linked fructose residues. Roots, sheaths, leaf blades and elongating leaves accumulated similar fructans although amounts of both low and high, and types of low, DP fructans, differed.• It is proposed that fructans in L. perenne are synthesized via four enzymes: 1-SST (1-sucrose-sucrosefructosyl transferase), 1-FFT (1-fructan-fructanfructosyl transferase), 6G-FT (6-glucose-fructosyl transferase) and 6-FFT (6-fructanfructanfructosyl transferase) or 6-SFT (6-sucrose-fructanfructosyl transferase).

The relative significance of the use of stored or currently absorbed C for the growth of leaves or roots of Lolium perenne L. after defoliation was assessed by steady‐state labelling of atmospheric CO2. Leaf growth for the first two days after defoliation was to a large extent dependent on the use of C reserves. The basal part of the elongating leaves was mainly new tissue and 91% of the C in this part of the leaf was derived from reserves assimilated prior to defoliation. However, half of the sucrose in the growth zone was produced from photosynthesis by the emerged leaves. Fructans that were initially present in elongating leaf bases were hydrolysed (loss of 93 to 100%) and the resulting fructose was found in the new leaf bases, suggesting that this pool may be used to support cell division and elongation. Despite a negative C balance at the whole‐plant level, fructans were synthesized from sucrose that was translocated to the new leaf bases. After a regrowth period of 28 d, 45% of the C fixed before defoliation was still present in the root and leaf tissue and only 1% was incorporated in entirely new tissue.

Summary• Longitudinal variability is reported in internal carbon partitioning as is the activity of enzymes involved in fructan synthesis during leaf development of Lolium perenne cv. Bravo.• Sink activity was reduced in L. perenne plants by cooling the roots while continuous illumination enhanced source activity, resulting in preferential accumulation of fructans at the leaf base. Fructan contents and enzymatic activity of two trisaccharides (SST (sucrose : sucrose fructosyltransferase) and 6G-FT (fructan : fructan 6G-fructosyltransferase) ) were measured in segments of elongating leaves, both mature leaf blades and sheaths.• The complement of fructans accumulated was similar in both elongating leaves and mature leaf sheaths. 6G-FT activity was found in L. perenne and characterized in a crude extract; this activity was greatest in the basal segment of elongating leaves and mature leaf sheaths. Throughout the leaf blade, however, SST and 6G-FT activities remained low and constant, and did not increase under conditions of sucrose accumulation.• Differences are shown in fructan accumulation between leaf tissues; the significance of these findings and the putative pathway of fructan synthesis in L. perenne are discussed.

Previous work on Lolium perenne showed that sucrose : sucrose fructosyltransferase (SST) activity did not increase concomitantly with fructan synthesis in regrowing leaves or in mature leaf blades of plants that have been subjected to carbohydrate-accumulating conditions. This was contrary to the pattern of SST activity in roots and stubble. To obtain further insight into the fructan synthesizing activities and to explain this discrepancy, total fructosyltransferase activity (FT) was assayed by increasing the sucrose and the soluble enzyme concentrations and was compared to sedimentable phlein sucrase activity (PS) throughout the regrowth period following defoliation in leaves, stubble and roots. Before analysis on 2-month-old plants, PS activity was characterized in dark-grown coleoptiles, using [U-"%C]sucrose. PS activity had a pH optimum of 6n0 and produced 1-kestose in addition to high molecular weight fructans with a mean DP of 9. In 2-month-old plants, sedimentable PS and FT soluble reactions contained an initial sucrose concentration of 160 mM and 400 mM and proteins equivalent to 1n4 and 2n1 g f. wt of tissue, respectively. In stubble and roots, the FT preparation catalysed the synthesis of large fructans, and the overall pattern resembled the native fructans when separated by anion exchange HPLC. In regrowing leaves, the FT preparation produced low-DP fructans relatively more than in vivo but synthesized the high-DP fructan characteristic of the tissue. Moreover, FT activity did not remain at a low level like SST activity but increased from day 5 after defoliation when fructans began to accumulate. PS activity formed very few low-DP fructans and 1-kestose was the main product. Trisaccharides generated by PS activity represented 2-5% of the total trisaccharide synthesis. High-DP fructans were detectable only when the products of the reaction were concentrated 100 times. Results are discussed with respect to the relative contribution of FT and PS activities for the synthesis of 1-kestose and fructans in roots, stubble and leaves of Lolium perenne.