Recipient Organization
UNIVERSITY OF MAINE
(N/A)
ORONO,ME 04469
Performing Department
(N/A)
Non Technical Summary
Lactic acid bacteria (LAB)-fermented foods have gained popularity in recent years as consumers learn more about their health-related benefits. Popular LAB-fermented foods consumed in the U.S. include pickles, olives, kimchi and sauerkraut. While fermented foods offer distinct health benefits, the elevated sodium content, which inhibits the growth of pathogenic and food spoilage organisms in these products, is a significant concern. Fermented food producers are investigating reduced-salt fermentation methods while seeking to maintain products free of foodborne pathogens and spoilage organisms and with low levels of allergenic amines. Expanding knowledge of vegetable fermentation processes will enhance food safety for producers in a developing niche, particularly those who market their products to consumers supporting farm-to-table, local, and organic agriculture. The National Agricultural Statistics Service (NASS) recorded a nearly 100% increase in the number of Maine farms producing fresh cabbage in the years between 2007 and 2012 (from 59 to 115). Ensuring the safety of value-added food products will also have a positive impact on the state's agricultural economy by providing fresh vegetable producers with knowledge and resources that support value-added production. We propose to ferment sauerkraut from locally produced cabbage, with natural bacterial flora and varied sodium content to assess acidification rates and the survival of inoculated pathogenic bacteria. Other analyses to be conducted include characterization and measurement of common biogenic amines that have negative health effects, and monitoring production of the healthful bioactive gamma-Aminobutyric acid (GABA) to improve nutritive value of fermented foods and the health of consumers.
Animal Health Component
90%
Research Effort Categories
Basic
(N/A)
Applied
90%
Developmental
10%
Goals / Objectives
OBJECTIVESEnhance Food Safety *Impact of reduced sodium concentration on pathogen survival during fermentationDescribe inactivation kinetics of foodborne pathogens during spontaneous vegetable fermentationObserve survival of pathogens at low concentrations during fermentation and cold storageMeasure titratable acidity (TA)Characterize organic acid profile by UPLC (lactic, acetic and other acids)Assess effect of salt concentration on formation of BABA concentrations during fermentation (by UPLC)BA concentrations during cold storage (3 months)BA content of fresh (regionally produced) and shelf-stable (canned) sauerkrautConfirm Product Acceptability *Consumer panels to determine acceptability of reduced-sodium products* Goals can still be accomplished with the reduced funding/time.
Project Methods
EXPERIMENTAL DESIGNPhase I: Raw, shredded cabbage purchased in bulk from a local distributor (to assure material homogeneity) will be analyzed for levels of common microbial indicator populations (TPC, lactic acid bacteria, coliforms, yeast and mold). Shredded cabbage will be combined with NaCl at 1.0, 1.5, 2.0 and 2.5% (w/w). All raw cabbage fermentations will be performed in triplicate, under sanitary conditions in 1 gallon fermentation vessels with airlocks and sample ports, maintained at 19° C. Because the proposed work will rely on natural cabbage microflora, the fermentation period will likely be longer than when using a known LAB inoculant. Initial fermentation may take 2 days to begin and several weeks to complete. During active fermentation, brine samples will be harvested from each vessel approximately every 72 hours with aliquots analyzed as follows:Salt concentration, pH and titratable acidity measurement;Identification and quantification of biogenic amines and organic acids with analytical methods modified for UPLC;Enumeration of aerobic microflora (dilution and plating on trypticase soy agar), yeast and mold (dilution and plating on acidified potato dextrose agar), lactic acid bacteria (dilution and plating on DeMann Rogosa Sharpe agar), and coliforms (most probable number analysis);Presence/absence of Listeria spp., Staphylococcus aureus and Salmonella spp. by enrichment and real time PCR;Presence/ absence of botulinum neurotoxin-producing clostridia determined by multiplex PCR.When brine pH reaches 3.4 (fermentation is complete), microbiological sampling will cease and fermentation vessels will be refrigerated at 4° C for 3 months. Samples will be harvested bi-weekly during the storage period, and tested for pH, TA, OA, BA.Phase II: Sauerkraut will be prepared as described previously, in one pint batches. Fermentation jars will be inoculated, upon the start of fermentation, with approximately 106 CFU/ml of pathogen cocktail comprised of three strains of enterohemorrhagic E. coli, Staphylococcus aureus, Salmonella, or Listeria monocytogenes. This phase will provide information regarding the susceptibility of the chosen pathogens to the conditions inherent in sauerkraut fermentation, and to clarify the effect of salt concentration on pathogen survival.Brine will be sampled for microbiological plating approximately every 72 hours throughout fermentation and biweekly for up to eight weeks of subsequent refrigerated storage at 4°C. Brine analysis will include:Aerobic plate count and LAB plate count to verify normal fermentative progress in the presence of pathogens;Count inoculated pathogens via dilution and plating on selective agar medium. Dilutions will be plated using non-selective agar overlay to allow for recovery and enumeration of injured cells.Plating will be discontinued when two successive samplings fail to produce counts above the detection limit of the method utilized (10 CFU/g). The pathogenic species with highest survivability during fermentation will be utilized in the final phase of research, as described below.Consumer acceptability of the control and reduced-sodium fermented products will be determined using the standard 9-point hedonic scale to assess appearance, taste, aroma, texture, and overall liking. One hundred adults (at least 18 years old) who are interested in reduced sodium foods and who like sauerkraut will be recruited from the greater Bangor, Maine region. SIMS software (version 6, Berkeley Heights, New Jersey) will be used to create the questionnaire, assign random 3-digit codes to samples, randomize sample presentation order, and analyze results. Samples will be served in 2-ounce white china ramekins at 4?°C. Spring water will be used to cleanse palates between samples. Consumers will record their liking on Hewlett Packard Elite Pro tablets. Demographic questions including age, gender, sauerkraut purchase habits, and purchase intent questions will also be asked. R software (The R Foundation for Statistical Computing, Vienna, Austria) will calculate analysis of variance, Tukey's HSD test, and preference mapping; a probability level of p ≤ 0.05 will be used to establish significant differences.Phase III: Sauerkraut will be prepared as described previously, with shredded cabbage inoculated with pathogen cell suspensions of approximately 102 CFU/jar to simulate low level contamination likely to occur naturally.Brine will be sampled for microbiological plating every 48 hours throughout fermentation and for up to eight weeks while in refrigerated storage at 4°C. Samples will be analyzed as follows:Aerobic plate count and LAB plate count to verify normal fermentative progression in the presence of pathogens;Presence or absence of inoculated pathogen by enrichment and cultural methods, without bacterial count.DATA ANALYSIS: Data will be analyzed using the statistical software packages SPSS. Organic acids, pH, titratable acidity, and salt concentrations will be correlated with pathogen survival/growth, and production of biogenic amines (BA).