Each head of lettuce was manually defoliated, and disposable
gloves, masks and caps were used during handling. Then, 25 g
portions were washed with sterile distilled water (250 mL) by
immersion in a previously sterilized container. In Sanitizing
Treatment 1 (S1), samples were washed using only sterile distilled
water, in S2 the samples were washed and immersed in 225 mL of
1% acetic acid for 15 min, and in S3 the samples were washed and
immersed in 225 mL of sodium hypochlorite at 150 mg L1 for
15 min. The control treatment consisted of the samples without any
type of sanitizing treatment.
The waste remaining from immersion in acetic acid (S2) was
neutralized in 225 mL of phosphate buffer (pH 7.0). To neutralize
the residues in samples that had been submitted to procedure S3,
2 mL of 10% sodium thiosulfate were added to the sodium hypochlorite
solution (Porto & Eiroa, 2006).
After the sanitization process, the fractions from each of the
treatments were individually homogenized in a sterile blender for
2 min at 2000 rpm with 225 mL of 0.1% peptone water. Each
homogenized portion was designated as the initial dilution (101)
of treatments, from which serial decimal dilutions were performed
(102e105) in the same diluent.