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ARS Home » Southeast Area » Raleigh, North Carolina » Food Science Research » Research » Research Project #428731

Research Project: Improved Processes for the Preservation and Utilization of Vegetables, Including Cucumber, Sweetpotato, Cabbage, and Peppers to Produce Safe, High Quality Products with Reduced Energy Use and Waste

Location: Food Science Research

2019 Annual Report

Objective 1. Enable the commercialization of reduced NaCl fermentation technology by identifying suitable starter cultures, optimizing processing parameters to prevent freezing of the fruits; and develop a shelf-life model that allows processors to rapidly and accurately predict end-use quality. Subobjective 1a: Develop starter cultures for cucumber fermentations brined with calcium chloride. Subobjective 1b: Identify parameters for long term storage of fermented cucumbers brined with calcium chloride so that freezing damage of the fruits can be prevented and removal of goods from tanks is facilitated during the winter season. Subobjective 1c: Develop an accelerated shelf-life model for fermented cucumber pickles. Objective 2. Determine the factors that maximize survival of probiotic bacteria and retention or production of healthful components in pickled vegetables to facilitate the commercialization of new cucumber pickling methods for delivery of live, probiotic, lactic acid bacteria to consumers. Subobjective 2a: Determine the acid resistance and growth characteristics of probiotic lactic acid bacteria that influence their survival in acidified and fermented cucumbers. Subobjective 2b: Identify suitable methods for the incorporation of probiotic lactic acid bacteria in refrigerated cucumber pickles. Subobjective 2c: Evaluate production and/or retention of bioactive peptides in fermented cucumbers. Objective 3. Enable new commercial methods for process-ready-fermentation or acidification to convert surplus vegetables at grocery stores and farmers' markets into value-added products with enhanced probiotic content. Objective 4. Integrate sweetpotato genotypes intended for commercial processing with their potential for increased levels of bioactive compounds and high quality food products.

Consumer interest in vegetable products with enhanced health benefits will drive future developments in the fermented and acidified vegetable industry. One major limitation for expansion is the generation of chloride waste and excess water use from traditional preservation technologies that rely on high sodium chloride (NaCl) concentrations. Cucumber fermentation in calcium chloride (CaCl2) brine was developed and transferred to commercial processors as an alternative to reduce chlorides in waste waters. Long-term adoption of this reduced NaCl preservation technology requires: a) defining conditions for bulk storage in outdoor tanks to prevent freezing damage in cold climates; b) understanding the microbial ecology of low salt vegetable fermentations so that appropriate starter cultures can be defined; and c) developing a shelf-life model to accurately predict end-use quality. The ability to deliver probiotic bacteria in pickles and to adapt acidification and fermentation technologies to preserve a wide range of surplus vegetables from farms and retail markets will provide new knowledge and technology for processors to engage the growing consumer market interested in healthy eating. For sweetpotatoes, consumer demands for readily available processed products have increased in recent years. Development of new cultivars that are bred for desirable processing characteristics, low acrylamide formation and high bioactive compound content, will benefit sweetpotato processors and farmers and boost the contribution of this nutritious vegetable in the U.S. diet. The proposed research will support developments that broaden the markets for sweetpotatoes, acidified and fermented vegetables and will reduce food and processing wastes.

Progress Report
Progress was made on all objectives which fall under NP306. Development of starter cultures for optimizing environmentally friendly low salt cucumber preservation by fermentation: This research aimed at optimizing the environmentally friendly low salt fermentations for commercial production of fermented vegetables. Starter cultures can accelerate the conversion of sugars while outcompeting the growth of undesired microorganisms. ARS researchers examined 243 potential lactic acid bacteria starter culture candidates isolated from commercial cucumber fermentations. ARS researchers selected 13 candidate starters based on a variety of tests, including utilization of cucumber sugars. None of these 13 starter culture candidates were found to produce potentially carcinogenic byproducts called biogenic amines. The starter culture candidates produced exopolysaccharides, which can be used as healthful prebiotics for human intestinal microbiota. Some of the selected starter culture candidates also produced antimicrobial compounds against Leuconostoc species, which are undesirable in commercial fermentations because they produce gas and contribute to the formation of hollow cavities (bloater defect) in fermented cucumbers. Antibiotic resistance profiling indicated the cultures were within the expected levels for this group of bacteria. The candidate starter cultures were, therefore, found to be suitable for use in commercial production of fermented cucumbers brined with low salt. Reduction of salt in bulk-stored fermented cucumber pickles: Fermented cucumbers are bulk-stored in cover brines containing 6 to 21% salt in outdoor tanks exposed to freezing temperatures in northern USA processing plants prior to the manufacture of hamburger pickle chips. Freeze and salting damage on fermented stocks was monitored by visually assessing external wrinkles and internal pits, instrumental texture analysis, colorimetric measurements, and determination of cucumber weight and water losses. A significant decrease in tissue firmness of fermented cucumbers was observed as a function of freezing, and a substantial visual difference was observed in internal tissue pittedness. A freezing damage appearance scale was developed to more accurately assess tissue pittedness. Freeze damage of fermented cucumbers was defined as loss in tissue firmness, darkening of peel color, development of pittedness and increased lactic acid efflux. Salting damage was defined as changes in skin color and shrinking of fermented cucumbers. This data will be used to help determine the need for salt levels in storage tanks during the winter season and reduce amounts of salt actually needed to retain quality and minimize production cost. Use of conserved DNA sequences to monitor cucumber processing: ARS researchers monitored the integrity of DNA as a tool to monitor cucumber processing steps typical of commercial products. Two techniques have been tested to monitor DNA integrity: the amplification of a non-nuclear (mitochondrial) gene and quantification of total cell DNA integrity. The analysis of fresh, fermented and pasteurized cucumbers suggested that significant degradation occurs to mitochondrial DNA due to heat processing and acidification. The total DNA integrity method was done with cucumber seeds as opposed to the fruit tissue DNA, but degradation was not detected in cucumbers exposed to blanching processes. While further research is needed, the methods developed may aid processers in analysis of processing parameters for fermented and acidified cucumber products. Market potential for probiotic pickles: Research to date shows great potential for the ability to deliver probiotic cultures in a refrigerated pickle product. A market study was conducted in collaboration with North Carolina State University to determine consumer attitudes towards novel, probiotic refrigerated pickle products. According to 1,100 consumers, price and brine cloudiness had the highest importance scores followed by package size/type and probiotic-specific label information. Consumers indicated that they would prefer pickles labeled as ‘Probiotic,’ and that ‘Supports digestive health’ was most preferred among the specific health-related label information. Consumers generally selected product characteristics to which they are accustomed (clear brines and similar in sour taste and spiciness to current products) although slightly cloudy brines such as those achieved through fermentation are likely to be accepted. These results show that there is an opportunity for probiotic pickles in the market and provides data on the product concepts that are preferred by consumers. Use of surplus vegetables (sweetpotatoes, broccoli, green peas, green beans, corn kernels, lettuce heads, tomatoes, green bell peppers, collard greens, and zucchini) from the United States market for value added fermented or acidified products and reduced waste: Conventional lactic acid fermentation was applied to pulverized, diced and whole vegetables that can be surplus at the farm and retail levels to determine the viability of the preservation technique for reducing food waste. Natural fermentations successfully preserved the vegetables. Addition of a starter culture, including probiotic cultures, to vegetable fermentations achieved equilibrium pH about 4-5 days faster than reliance on the natural microbiota. A combination of blanching and a starter culture supplement also reduced the fermentation time. Preservation of surplus vegetables without fermentation by acidification was effective at pH 3.4 ± 0.1 with vinegar and natural preservatives in closed jars. These studies indicate that lactic acid fermentation and/or acidification methods could be refined into a simple procedure for preservation of a wide range of surplus produce to help satisfy global demand for nutritious food. Whole cucumber blanching for improving the quality of refrigerated pickles: Refrigerated pickle quality relies primarily on a fresh cucumber-like appearance and crisp, crunchy texture. Changes in appearance and texture during one year of storage were determined for refrigerated pickles prepared from fresh cucumbers or cucumbers blanched in hot (80°C) water prior to cutting into spears and pickling. Whole cucumber blanching for 90 to 180 sec consistently reduced naturally occurring bacteria by at least 100 fold, and would provide sufficient reduction in pathogens on the cucumber surface to help meet FDA standards. Blanching assisted in maintaining the color of the cucumber peels, and retarded cure appearance development. This research contributes to the development of improved processes for the preservation of cucumbers to improve the safety and quality of consumer products. Identification of health-promoting amino acids in fermented vegetables: While much is known about the content of essential amino acids in fresh cucumbers, there is limited information available on bioactive amino acids which may impact human health or physiology and on their changes during cucumber preservation. Gamma-amino butyric acid (GABA) is a non-protein amino acid that may be present in both fresh and fermented foods and has been shown to promote antihypertensive and antianxiety effects and improve cognitive function. A mass spectrometry based method was developed to quantify GABA and other amino acids in raw, acidified, and fermented cucumbers. Experiments showed that the content of several amino acids differed between acidified and fermented cucumbers, including production of GABA during cucumber fermentation. This research contributes to our understanding of changes in health-promoting compounds that occur during lactic acid fermentation of vegetables. Reducing acrylamide in fried sweetpotato products through rapid detection methods and chemical knowledge to facilitate new variety development. Acrylamide, a known neurotoxin and potential carcinogen, can be formed in carbohydrate rich foods including sweetpotato chips and french fries. Research was conducted to develop a rapid method for estimating the acrylamide content in sweetpotato chips using near-infrared spectroscopy. The method can be used to screen hundreds of samples without the need for organic solvents and expensive analytical instruments. The data showed that the models used with white potatos are not sufficient to predict acrylamide formation in sweetpotato matrix. This research showed that the chemical composition of sweetpotato genotypes is an important selection criteria for development of new sweetpotato varieties adapted for fried products with low acrylamide formation. Sweetpotato French-fry texture: Several sweet potato cultivars that ranged in dry matter content were analyzed for chemical composition and sensory texture characteristics for production of French fries. Starch pasting properties were examined to better understand the components of dry matter that control the texture of sweetpotato French-fries. Sweetpotato starch characteristics of 16 varieties correlated density and moisture of fries. These results suggest that further work on sweetpotato varieties with specific starch profiles could be used to help achieve consumer-preferred texture attributes. Evaluation of sweetpotato breeding lines: Chips were prepared from 61 clones in the North Carolina State breeding lines resulting in the selection of 17 clones for further breeding cycles and evaluation. French-fries from 79 sweetpotato clones were prepared for sensory screening with the goal of selecting 16-20 clones for further work. Additionally, sugar profiles were quantified in raw and baked sweetpotatoes for ~120 clones from a large genetic mapping population to assist in determining the association between quality traits of raw and baked storage roots at harvest and during post-harvest storage.

1. Strategies to mitigate freezing damage on bulk stored fermented cucumbers. Cucumbers are commercially fermented for the manufacture of pickles and stored in bulk in outdoor open top fiberglass tanks. During winter, snow and ice that accumulate around and on top of the tanks influence heat transfer in an unpredictable manner, often compromising the quality of the fermented cucumbers due to freezing. The capacity of inexpensive and resilient fermentation tank insulation to prevent freezing of the fermentation cover brine located near the tank walls and on the surface was evaluated by ARS scientists in Raleigh, North Carolina. Adding insulation on top of the fermentation tanks in the form of a cover significantly reduce the freezing of the fermented fruits and cover brine located at the top of the tanks at 32°F for 4 days. Additionally, we found that the use of 14.5% glycerin, 18% calcium chloride or 14% glycerin and 5% calcium chloride can reduce the freezing of cover brines and the formation of ice. These findings aid in reducing freeze induced damage on fermented cucumbers and provide alternative strategies to achieve the reduction of the formation of a thick layer of ice on top of fermentation tanks during the winter in the Northern United States.

2. Effects of brine acidification on cucumber fermentations in calcium or sodium chloride brines. Commercial fermentation for bulk preservation of cucumbers typically relies on natural microbiota and brines with high concentrations of sodium chloride (NaCl). An alternative process utilizing low salt calcium chloride brines was previously developed to eliminate NaCl from fermentation brines for reduced environmental impact. In this work, ARS scientists in Raleigh, North Carolina, studied the effects of initial brine acidification on the course of lactic acid fermentation and resulting texture quality of cucumbers fermented in calcium or sodium salt brines. Fermentation brines containing acetic acid (the acid in vinegar) reduced the pH of the cucumber and the soil-associated Enterobacteriaceae spp. most rapidly and favored the conversion of sugars to lactic acid. Interestingly, the texture quality was not affected by brine acidification, and all cucumbers fermented in calcium brines in the absence of air retained their firmness during fermentation and bulk storage, further demonstrating the suitability of this fermentation technology to produce high quality products with reduced environmental impact under controlled conditions.

3. Pickle peptides. Health-promoting compounds composed of particular short chains of amino acids are known as bioactive peptides. Bioactive peptides possessing therapeutic properties are well documented in foods fermented by lactic acid bacteria, including dairy products, sourdough, and cured meats, but have not been previously investigated in fermented vegetables. Cucumber pickles, the most commonly consumed fermented vegetable in the United States, are not amenable to analytical workflows used for the analysis of peptides without extensive sample preparation, due to their high salt content. A novel mass spectrometry-based method was used by ARS scientists in Raleigh, North Carolina, for direct analysis of peptides in acidified and fermented cucumbers. Five short peptides with known anti-hypertensive properties were identified in fermented cucumber: lysine-proline, isoleucine-proline-proline, leucine-proline-proline, valine-proline-proline, and arginine-tyrosine. The results showed that four of the five bioactive peptides were uniquely present in fermented cucumber. Bioactive peptides, although present at low concentrations compared to other food constituents, may be valuable functional food components due to their high potency. Further research is needed to determine if these compositional changes during lactic acid fermentation are in sufficient quantities for a health benefit.

Review Publications
Perez Diaz, I.M., Hayes, J.S., Medina, E., Webber, A., Butz, N., Dickey, A., Lu, Z., Azcarate-Peril, M.A. 2019. Assessment of the non-lactic acid bacteria microbiota in fresh cucumbers and commercially fermented cucumber pickles brined with 6% NaCl. Food Microbiology. 77:10-20.
Fideler, J., Johanningsmeier, S.D., Ekelof, M., Muddiman, D.C. 2019. Discovery and quantification of bioactive peptides in fermented cucumber by direct analysis IR-MALDESI mass spectrometry and LC-QQQ-MS. Journal of Food Chemistry. 271:715-723.
Klevorn, C.M., Dean, L.L., Johanningsmeier, S.D. 2019. Metabolite profiles of raw peanut seeds reveal differences between market-types. Journal of Food Science. 84(3):397-405.
McMurtrie, E.K., Johanningsmeier, S.D., Price, R.E., Breidt, F. 2019. Effect of brine acidification on fermentation microbiota and texture quality of cucumbers fermented in calcium chloride brines. Journal of Food Science. 84(5):1129-1137.