Of the many responses of plants to elevated CO2, accumulation of total non-structural carbohydrates (TNC in % dry weight) in leaves is one of the most consistent. Insufficient sink activity or transport capacity may explain this obvious disparity between CO2 assimilation and carbohydrate dissipation and structural investment. If transport capacity contributes to the problem, phloem loading may be the crucial step. It has been hypothesized that symplastic phloem loading is less efficient than apoplastic phloem loading, and hence plant species using the symplastic pathway and growing under high light and good water supply should accumulate more TNC at any given CO2 level, but particularly under elevated CO2. We tested this hypothesis by carrying out CO2 enrichment experiments with 28 plant species known to belong to groups of contrasting phloem-loading type. Under current ambient CO2 symplastic loaders were found to accumulate 36% TNC compared with only 19% in apoplastic loaders (P=0.0016). CO2 enrichment to 600 μmol mol−1 increased TNC in both groups by the same absolute amount, bringing the mean TNC level to 41% in symplastic loaders (compared to 25% in apoplastic loaders), which may be close to TNC saturation (coupled with chlornplast malfunction). Eight tree species, ranked as symplastic loaders by their minor vein companion cell configuration, showed TNC responses more similar to those of apoplastic herbaceous loaders. Similar results are obtained when TNC is expressed on a unit leaf area basis, since mean specific leaf areas of groups were not significantly different. We conclude that phloem loading has a surprisingly strong effect on leaf tissue composition, and thus may translate into alterations of food webs and ecosystem functioning, particularly under high CO2.