Rubber trees (Hevea brasiliensis) are the main source of natural rubber, extracted from latex, which exudes from the trunk after tapping. Tapped trees require large amounts of carbon (C) to regenerate the latex after its collection. Knowing the contribution of C sources involved in latex biosynthesis will help in understanding how rubber trees face this additional C demand. Whole crown 13CO2 pulse labelling was performed on 4-year-old rubber trees in June, when latex production was low, and in October, when it was high. 13C content was quantified in the foliage, phloem sap, wood, and latex. In both labelling periods, 13C was recovered in latex just after labelling, indicating that part of the carbohydrate was directly allocated to latex. However, significant amounts of 13C were still recovered in latex after 100 d and the peak was reached significantly later than in phloem sap, demonstrating the contribution of a reserve pool as a source of latex C. The contribution of new photosynthates to latex regeneration was faster and higher when latex metabolism was well established, in October, than in June. An improved understanding of C dynamics and the source–sink relationship in rubber tree is crucial to adapt tapping system practices and ensure sustainable latex production.