Bentonite is mined globally for use in commercial and industrial applications. In these applications, smectite content and composition are the paramount factors of the bentonite material and control its properties. As bentonite composition and properties can vary significantly over a large mining district or within a single mine, quality control is required including: mineral composition, especially smectite content; cation exchange capacity (CEC); exchangeable cation composition; and smectite crystallochemical features. Differences in bentonite composition locally or over a spatial area stem from the different geological settings present throughout bentonitization. The study aims were to: (1) determine the layer charge (LC) variation of dioctahedral smectite over the Bavarian mining district and within individual mines in the area; and (2) assess the error in smectite content calculations based on CEC data resulting from the actual range of experimentally determined LC values. This information has been missing in the scientific literature, as previous LC methods were laborious or subject to assumptions, making a comprehensive study over a large spatial area impractical. This study employed the use of the recently developed efficient and precise spectroscopic ‘O-D method’, which enabled the LC measurement of 40 samples from eight mines in the Bavarian bentonite mining district, covering an area of 250 km2, within the North Alpine Foreland Basin. Results showed LC values calibrated against the alkylammonium method (LC (AAM)) generally ranged between 0.29 and 0.30 eq per formula unit (FU), with only 10% of samples showing LC values >0.31 eq/FU. This narrow LC range has positive implications for the accuracy of determining smectite content calculated from CEC data, during routine quality control of Bavarian and other bentonites. The average error of the CEC-based smectite contents resulting from LC variations was, on average, ±3 wt.%.