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The efficacy of various solutions of Pro-SanTM were tested on romaine lettuce leaves against Salmonella enterica. Romaine lettuce samples were inoculated with a 5-strain cocktail of Nalidixic acid resistant Salmonella enterica and held at room temperature for 16-18 hours to allow a biofilm to form. Lettuce samples were then sprayed separately with one of six treatments and remained exposed to the treatment for various times (including 30, 45, 60, 75, 90, 105, 120, 280, and 240 minutes) which was at either refrigerated temperature or room temperature. Samples were then plated onto nalidixic acid supplemented Xylose Lysine Desoxycholate (XLD) agar and bacterial colonies were enumerated 48 hours later after being held at 35° C. The 6 spray treatments included: a distilled water control, a 150 ppm chlorine bleach solution (with pH maintained between 6.4 and 7.5), a 0.78% Pro-SanTM LC solution, a 0.19% Pro-SanTM LC solution, a 0.78% Pro-SanTM LC Soft solution, and a 0.19% Pro-SanTM LC Soft solution. Samples of lettuce leaves that were not sprayed served as dry controls. Pro-SanTM LC and Pro-SanTM LC Soft are both bio-degradable GRAS sanitizers composed of an organic acid and a surfactant, with a pH around 2.2. After 3 replicates at each application time, data was analyzed using SAS software. Three variables were analyzed: time, treatment, and treatment*time (to determine if there is any interaction between the two variables). The time and treatment variables had significant 36 differences (a among the treatments was not different among the times. Therefore, Salmonella counts for each time were logged and averaged. A Tukey pairwise t-test was obtained from this data, which indicated that the dry control, distilled water control, and 150 ppm bleach solution were not significantly different from each other, but were less effective than the Pro-SanTM solutions when the treatments were tested at room temperature. At refrigerated temperature, the same trend among treatments existed but only the 0.78% solutions were statistically more effective than the dry control and distilled water control. Additionally, only the 0.78% Pro-SanTM was more effective than the bleach solution. The ability of the increased concentrations of Pro-SanTM LC to kill bacteria present on lettuce leaves is theorized to be due to the ability of the surfactant to aid the delivery of the primary antimicrobial agent (the organic acid) to the cells by penetrating the biofilm (although the surfactant itself also has some antimicrobial activity) (Feliciano 2009). Furthermore, it is hypothesized that when a biofilm is present, the organic matter inactivates the majority of the hypochlorite ion in the bleach solution, making it ineffective in this condition. The effects of each spray treatment on the color of the lettuce leaves was also analyzed with a Hunter L*a*b* system, and data showed that the color was not significantly different among any treatment groups. Overall quality was also not effected either treatment, based on results of a triangle tests completed with a total of 100 panelists.

Inactivation of Salmonella enterica on romaine lettuce following spraying with Pro-San<sup>TM</sup> - a biodegradable foodgrade sanitizer