Background: National forest inventories (NFI) have a long history providing data to obtain nationally representative and accurate estimates of growing stock. Today, in most NFIs additional data are collected to provide information on a range of forest ecosystem functions such as biodiversity, habitat, nutrient and carbon dynamics. An important driver of nutrient and C cycling is decomposing biomass produced by forest vegetation. Several studies have demonstrated that understory vegetation, particularly annual plant litter of the herb layer can contribute significantly to nutrient and C cycling in forests. A methodology to obtain comprehensive, consistent and nationally representative estimates of herb layer biomass on NFI plots could provide added value to NFIs by complementing the existing strong basis of biomass estimates of the tree and tall shrub layer. The study was based on data from the Swiss NFI since it covers a large environmental gradient, which extends its applicability to other NFIs. Results: Based on data from 405 measurements in nine forest strata, a parsimonious model formulation was identified to predict total and non-ligneous herb layer biomass. Besides herb layer cover, elevation was the main statistically significant explanatory variable for biomass. The regression models accurately predicted biomass based on absolute percentage cover (for total biomass: R 2 = 0.65, p = 0; for non-ligneous biomass: R 2 = 0.76; p = 0) as well as on cover classes (R 2 = 0.83; p = 0; and R 2 = 0.79, p = 0), which are typically used in NFIs. The good performance was supported by the verification with data from repeated samples. For the 2nd, 3rd, and 4th Swiss NFI estimates of non-ligneous above-ground herb layer biomass 586.6 ± 7.7, 575.2 ± 7.6, and 586.7 ± 7.9 kg•ha − 1 , respectively. Conclusions: The study presents a methodology to obtain herb layer biomass estimates based on a harmonized and standardized attribute available in many NFIs. The result of this study was a parsimonious model requiring only elevation data of sample plots in addition to NFI cover estimates to provide unbiased estimates at the national scale. These qualities are particularly important as they ensure accurate, consistent, and comparable results.