Cocoa pods (Trinitario variety) were harvested from plantations in
North Queensland, Australia and transported to UNSW Australia Sydney where they were stored at 20–25 °C and used within
7–10 days from harvest. The pods were then split for manual removal
of the beans. The mass of beans was mixed on a tray and deliberately
brought into contact with the outer surfaces of extracted pods to provide
a source of natural microbial inoculum. The beanmasswas then divided
into two ×5 kg batches which were transferred into two plastic
boxes (17 × 17 × 20 cm) for fermentation. To restrict bacterial growth,
a solution of Nisaplin (Danisco, Denmark) containing 2.5% of pure nisin
or amixture of nisin and lysozyme (Delvozyme, DSM, The Netherlands)
was sprayed onto the beans in one box and mixed to give uniform distribution
of nisin and lysozyme. The cocoa beans in the other box
were not treated with nisin or lysozyme but similarly mixed. The
boxes of beans were then incubated for fermentation which developed
spontaneously due to growth of the indigenous microbiota