Grantee Research Project Results
Final Report: Novel Biopreservatives to Improve the Shelf Life of Dairy Products
EPA Contract Number: 68HERC22C0018Title: Novel Biopreservatives to Improve the Shelf Life of Dairy Products
Investigators: Sheth, Ravi
Small Business: Imvela Corp
EPA Contact: Richards, April
Phase: I
Project Period: December 1, 2021 through May 31, 2022
Project Amount: $99,309
RFA: Small Business Innovation Research (SBIR) Phase I (2022) RFA Text | Recipients Lists
Research Category: Small Business Innovation Research (SBIR)
Description:
Overview: In this Phase I SBIR project, supported by the EPA SBIR Program under contract number 68HERC22C0018, Imvela Corp. started development of a natural microbial bioprotectant for dairy products, with the goal of decreasing food waste and the associated environmental impacts (including greenhouse gasses and water and energy use). Imvela Corp. started development focused on cottage cheese products, which are widely consumed in the US (more than 600 million pounds produced in the US annually) and have a relatively short shelf-life. Imvela Corp. worked closely with a commercial partner, under contract to purchase the solution once developed, that has a top-selling line of cottage cheese products already available for sale across the US, to ensure the bioprotectant meets real-world customer standards. The core technology developed through this work can be broadened to design bioprotectant products applicable to other products beyond cottage cheese, starting with other dairy products and eventually other types of ready-to-eat food. The Phase I work was intended to lay the technical and commercial foundations for additional follow-on work that can result in a validated product in the marketplace within a year of additional work.
Phase I research activities were geared towards identifying and assembling novel microbial communities optimized to be bioprotectants against cottage cheese spoilage organisms. We identified primary spoilage organisms (SOs) from real cottage cheese products, developed high content/high throughput microscopy screening systems to quantify microorganisms (the SOs) in a matrix with customized growth media to approximate cottage cheese, and developed assays to replicate spoilage conditions. We conducted high volume screens of microorganisms, individually and in combination in our biobank against a SO, and compared the inhibition of growth that results against two competitor bioprotectant products.
This work has high practical and commercial potential, starting with the cottage cheese market and as demonstrated by our customer making a firm commitment to purchase the solution we develop. Existing bioprotection options on the market today (e.g. naturally produced propionic acid or natural bacteriocins) do not meet the high performance requirements or parity with chemical preservatives required by many consumer brands. Consumer preferences for clean-label products featuring natural, non synthetic, and GMO-free solutions, combined with manufacturers’ desire to extend shelf life and reduce costly waste, drive demand for improved bioprotection approaches. While our beachhead market is cottage cheese (~$1.1B in US sales, which could drive several million dollars in annual revenue for our solution), we anticipate working to expand the biopreservative solution into adjacent markets in the dairy space such as other soft cheese, sour cream, liquid dairy, and yogurt. Dairy products are the second largest category of wasted food (after fruits and vegetables), according to the EPA, indicating the significant commercial potential and opportunity for positive environmental and social impacts from better biopreservative solutions in the space.
Purpose: The key problem we’re addressing is inhibiting the growth of spoilage organisms (SOs) in cottage cheese, thereby reducing food waste and its associated environmental impacts. We will produce a microbial-based product that improves cottage cheese shelf-life compared to other biopreservatives currently on the market by exploring a larger, more diverse microbial space and implementing a high throughput, high resolution microscopy-based screen and computational methods. We will produce a Superculture Ingredient (SCI) from a consortia of microbes that have antifungal properties.
Summary/Accomplishments (Outputs/Outcomes):
Purpose: The key problem we’re addressing is inhibiting the growth of spoilage organisms (SOs) in cottage cheese, thereby reducing food waste and its associated environmental impacts. We will produce a microbial-based product that improves cottage cheese shelf-life compared to other biopreservatives currently on the market by exploring a larger, more diverse microbial space and implementing a high throughput, high resolution microscopy-based screen and computational methods. We will produce a Superculture Ingredient (SCI) from a consortia of microbes that have antifungal properties.
Work Carried Out and Results: We biobanked >50 “spoiled” cottage cheese samples across 5 brands and utilized a state of the art high throughput colony isolation platform to perform isolation of samples. We then included one of these spoilage organisms in our screen. We onboarded other relevant SOs from both external sources and our biobank. These SOs were based on either customer reports from manufacturing plants or primary literature. We identified 7 potential SOs for the screen and, as we are focusing on fungi, we have included examples of both yeast and mold. To ensure the success of our screen we characterized the growth dynamics of target SOs in standard growth media using the accelerated growth conditions needed for screening. Based on these experiments we chose 5 target SOs for our initial screen.
We have also established the appropriate substrate for our screen. We attempted to image SOs in the context of primary material (diluted cottage cheese) and found that this method has poor signal-to-noise compared to optically transparent growth media. Thus, we generated a custom-designed, cottage-cheese-like, optically transparent media. Additionally, we found that current biopreservative solutions did not inhibit yeast growth under our screening conditions. Thus, we tested chemical preservatives used in cottage cheese including lactic acid, citric acid, and propionic acid, as positive controls for growth inhibition and found that propionic acid was a strong inhibitor of growth of multiple SOs under our conditions.
We generated fermentates for our screen by fermenting individual microbes from our internal biobank. We then implemented a combination of custom Python scripts and automation robotics software to effectively build screening plates. Lastly, we optimized an imaging protocol which includes staining cells as needed, fixing samples, performing serial dilution, and imaging samples with brightfield or widefield fluorescence modalities, depending on the specific SO (Figure 1).
Figure 1 - Imaging methodology and modality have been optimized for our target SOs. Examples of Kluyveromyces marxianus (A, fluorescence with DAPI nuclear staining), and Clavispora lusitaniae (B, brightfield).
Our preliminary results suggest that a subset of putative bioprotectants from our screen inhibit SO growth compared to vehicle controls, a condition that was not fulfilled by a product currently used by our commercial partner. We also have data from a related product that disrupts biofilms indicating that SCIs have increased efficacy compared to single strains (Figure 2).
Figure 2 - SCIs increase disruption of biofilms compared to single microbial strains. Vehicle control (black), positive control for disruption (red), fermentates from single microbial strains (blue), and SCIs of fermentates from a consortia of microbes (green) are shown. Solid lines indicate the estimated marginal mean of the treatment group, dashed lines indicate the 95% confidence intervals.
Conclusions:
In Phase I, we have established all the methodology necessary to screen for novel bioprotectants against at least 5 target SOs using high throughput microscopy and generated an appropriate cottage-cheese-like matrix. We have begun our screen and have preliminary data suggesting that putative bioprotectants from the screen inhibit SO growth and that communities of microbial ingredients have greater efficacy than solutions generated by single microbial strains. This gives us confidence that we will be able to generate a natural bioprotectant solution that outperforms existing solutions on the market, and contribute to a significant reduction in food waste in cottage cheese products. The research foundation will also allow us to extend the technology’s application to other varieties of dairy products.
Commercialization Potential: The work supported by the EPA Phase I SBIR award was designed from the start to have high and direct practical and commercial potential, starting with the cottage cheese market and expanding to other dairy categories. The cottage cheese market alone reached $1.1B in US sales in 2021, according to market research firm IRI, which we believe could drive at least $5M in annual revenue for our solution based on initial conversations with prospective customers. Once we have established a working biopreservative solution for cottage cheese, we intend to extend the solution to adjacent markets. The overall global dairy market is very large, at ~$770B per year with ~5% annual growth, and offers substantial market opportunity.
Throughout the Phase I period, we have worked closely with an anchor customer, a leading cottage cheese manufacturer in the US, which has made firm commitments to purchase the solution and apply it to their products. Assuming technical progress continues as planned, we expect that our customer will begin purchasing and manufacturing products with our solution mid-2023 for a full-scale production and commercial test, transitioning into full-scale supply later that year.
We have also validated the commercial potential of this technology through conversations with other prospective customers. Dairy producers value ways of extending the shelf life of their products because they believe it can increase sales (by reducing stockouts at retail and increasing value to consumers), reduce costs (by decreasing the amount of product wasted or returned from the supply chain), and help them meet their social responsibility targets (by reducing food waste and the associated environmental impacts). Further, these customers have clearly stated that end consumers are increasingly aware of the ingredient label and prefer products with natural ingredients and that are free of artificial chemicals. Existing solutions in the marketplace cannot fully satisfy both of these demands.
Market research confirms the significant opportunity available in this area. The dairy market is large and growing– more than $770B of dairy products are sold globally every year and sales are growing at ~5% per year– while food waste in the sector is high. The EPA’s 2021 report on the Environmental Impacts of US Food Waste indicates that dairy is the second most wasted category of food (after fruits and vegetables), and among food types is one of the largest users of water and energy. According to the International Dairy Federation, about one fifth of all dairy products get wasted, and 55% of that waste happens on the consumer side. Extending shelf life of dairy products is an important mechanism that could help significantly reduce this waste and its associated environmental impacts.
Our biopreservative solution will offer important advantages over existing bioprotection options on the market today. Compared to incumbent natural protection options, such as naturally produced propionic acid or natural bacteriocins, our solution will offer significantly enhanced protective performance, thereby extending shelf life and reducing costly food waste. Compared to traditional chemical-based biopreservatives, our solution will allow food manufacturers to better meet consumer preferences for clean-label products.
A key element of our commercialization plan is for us to retain the intellectual property to the solution, which will allow us to file for a patent and sell the solution to other producers in the broader dairy industry. The technical groundwork we lay during this project will support future work that can go beyond cottage cheese and even dairy at large. In the future, we believe we can apply our platform to develop biopreservative solutions in the other top loss categories as well, including fresh fruits and vegetables (19.6% of all spoiled food); processed fruit and vegetables (8.6% of all spoiled food); and meat, poultry, and fish (8.5% of all spoiled food).
We will apply our experience in successfully commercializing other products to bring this dairy bioprotectant solution to market. As described above, we have built a strong relationship with our anchor customer to ensure a very high likelihood that the solution will be commercially adopted and accelerate its time to market. Working closely with a customer ensures that our solution adequately addresses real-world issues and can be incorporated into large-scale manufacturing processes that produce millions of pounds of product a year. Further, our existing customer contract provides key commercial inputs, such as the ingredient price, that will make entering into a supply agreement faster than if we were dealing with a new customer.
To product the dairy bioprotectant solution, we anticipate purchasing most component strains from commodity suppliers and contracting with contract suppliers for the production of up to 2 specialty strains. We will conduct quality testing, blending, packaging, and order fulfillment in-house, using the same general process that we developed for prior products. In addition to funding SBIR funding from the EPA, we will fund our commercialization efforts with revenue from customers and from funds we have already raised from investors (we raised a Series A round from private-sector investors in late 2021).
The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.