The aim of this study was to investigate the prediction of 14C-phenanthrene and 14C-hexadecane biodegradation in the presence of other hydrocarbons in soil using β- and α-cyclodextrin (CD) solutions, respectively. Prediction of the biodegradation of 14C-phenanthrene using the β-CD extraction was robust under single, co-contaminant and multiple contaminant conditions (r2 = 0.92, slope of best fit line = 0.87, intercept = 7.24, n = 84). Prediction of 14C-hexadecane using the α-CD extraction was robust under single and co-contaminant conditions (r2 = 0.92, slope of best fit line = 0.97, intercept = 1.24, n = 60); however, the α-CD could not accurately predict 14C-hexadecane biodegradation in the presence of multiple contaminants. The presence of multiple contaminants enhanced 14C-hexadecane mineralisation, but did not enhance extractability. The results from this study provide further evidence for the application of HPCD extractions for the measurement of microbial accessibility in soil.