We present an 8000-year biomarker and stable carbon isotope record from the Verlorenvlei Estuary, South Africa. We assessed how leaf wax lipids, insoluble macromolecular organic matter, bulk C/N data and compound-specific stable carbon isotopes were linked to the site’s palynological record and to evidence for regional sea level and environmental change. Down-core trends in bulk δ13C are closely coupled to trends in pollen types from saline-tolerant taxa. These trends are mirrored by variations in the incorporation of reduced sulphur into macromolecular organic matter. This process, quantified with the thiophene ratio, is closely associated with periods of higher sea level 8,000–4,300 cal yr BP. We propose the thiophene ratio is a proxy for relative marine influence within (peri) estuarine sediments. All measured variables indicate differences between early-middle Holocene (8,000–4,300 cal BP) and late Holocene conditions at Verlorenvlei. The former period was more saline and preserves more labile macromolecular organic matter. Marine influence declined after 4,300 cal yr BP, and although the abundance of short-chain-length n-alkanes suggests continued presence of wetland flora until 2,500 cal yr BP, organic matter preservation became poorer and a drying trend was inferred, most notably for the interval 2,500–900 cal BP. Increasing freshwater inundation is apparent during the last 700 cal yr, consistent with several records from this region. Leaf wax n-alkane distributions are largely uncorrelated with bulk organic matter variables, with the exception of the abundance of C31 and C33 n-alkanes, which are negatively correlated with δ13CTOC. Furthermore, C31–C33 n-alkane δ13C values are uncorrelated with C23–C29 δ13C and δ13CTOC. They are also higher than our newly measured terrestrial (C3) vegetation C29 and C31 end-member values of −35 ± 2 and −34 ± 1 ‰, respectively. These patterns are best explained by a dominant contribution of local riparian vegetation to the C23–C29 n-alkanes, but time-varying contributions of non-local leaf waxes to the C31–C33 signals. This renders inferences concerning regional environmental change from long-chain leaf waxes potentially challenging in this setting.