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Yarrowia lipolytica and Lactobacillus paracasei Solid State Fermentation as a Valuable Biotechnological Tool for the Pork Lard and Okara's Biotransformation.

ABSTRACT: This study reports the biovalorization of the two agri-food by-products (pork lard and freeze-dried okara) through solid-state fermentation using a monoculture of Yarrowia lipolytica or a co-culture of Y. lipolytica and Lactobacillus paracasei, for developing a valuable fermented product with antioxidant and antimicrobial activity. First, some yeast strains were selected based on their properties to produce enzymes (protease and lipase) by cultivation on 5% (w/v) pork lard or 2% (w/v) freeze-dried okara. Two selected strains, Y. lipolytica MIUG D5 and Y. lipolytica ATCC 18942, were further used for the fermentation alone or in a co-culture with L. paracasei MIUG BL2. The Plackett-Burman experimental design was used to establish the effects of the fermentation parameters in order to obtain a fermented product with improved antioxidant and antimicrobial activities. As the Plackett-Burman experimental design are independent variables, the concentrations of the freeze-dried okara, pork lard, glycerol, inoculums type, inoculum concentration, and the fermentation time were analyzed. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging potential and the antimicrobial activity against aerobic spore-forming microorganisms were assessed as responses. For the fermented products, an antioxidant potential between 6.77-17.78 mM TE/g was obtained while the antimicrobial activity against Aspergillus niger ranged from 24 to 64%. Based on the statistical analysis, the time of the yeast fermentation and the concentration of pork lard were selected as variables with the influence on the SSF fermentation process and the functional properties of the fermented product. In the general context of a circular economy, the results demonstrate the possibility of bio-transforming the freeze-dried okara and the pork lard using Y. lipolytica as a valuable workhorse for the lactic acid bacteria (LAB) metabolism and postbiotics production into a fermented product, which is recommended for use as a food and feed ingredient with biotic properties.

SUBMITTER: Cotarlet M

PROVIDER: S-EPMC7464363 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Yarrowia lipolytica and Lactobacillus paracasei Solid State Fermentation as a Valuable Biotechnological Tool for the Pork Lard and Okara's Biotransformation.

Cotârleț Mihaela M Stănciuc Nicoleta N Bahrim Gabriela Elena GE

Microorganisms 20200722 8

This study reports the biovalorization of the two agri-food by-products (pork lard and freeze-dried okara) through solid-state fermentation using a monoculture of Yarrowia lipolytica or a co-culture of Y. lipolytica and Lactobacillus paracasei, for developing a valuable fermented product with antioxidant and antimicrobial activity. First, some yeast strains were selected based on their properties to produce enzymes (protease and lipase) by cultivation on 5% (w/v</i ...[more]

PMID: 32708033

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Project description:The yeast strain Yarrowia lipolytica IPS 21 was tested for its ability to degrade potentially toxic chrome-tanned leather shavings (CTLS) in a liquid environment. Biological and chemical parameters were monitored during a 48-h period of biotransformation of the protein-rich waste. CTLS was added at a concentration of 0.1-4% (wt/wt) to a modified YPG medium (15 g L-1 yeast extract and 5 g L-1 NaCl). Biodegradation and bioconversion were performed in a one-step process. It was found that the higher degradation rate depended on the activity of the proteases and the pH of the medium, but not on the initial inoculum ratio and the activity of the dehydrogenase. The highest efficiency of the process was obtained for 4% (wt/wt) CTLS on day 2 (degradation rate 58-67%, biomass production 2.11-2.20 g L-1, protease activity 312 U mg-1 protein, and pH 9.20). Our results showed that total chromium was probably not transported across the cytoplasmic membrane of Y. lipolytica IPS21 and that chromium (III) was not oxidized to chromium (VI). The phytotoxicity of selected amino acid supernatants [2.5% (vol/vol)] was tested after the bioconversion process. It was found that the supernatants had a stimulating effect on the plants tested. The root elongation was 29-28% higher than that of the reference samples. This result makes Y. lipolytica IPS21 a potential candidate for safely converting potentially toxic protein-rich wastes into valuable products without enzyme isolation, e.g., amino acid fertilizers. IMPORTANCE Enzyme technologies have the greatest practical relevance to environmental trends. Overcoming the barrier of the high cost of carbon substrates used for biotransformation is the main challenge of these methods. The huge potential of the use of extracellular proteases of Yarrowia species or amino acids in various industries indicates the need for the extension of basic research on waste as a carbon source for this yeast. The experiments demonstrated that it is possible to use Y. lipolytica IPS21 for bioconversion of chrome-tanned leather shavings (CTLS) in a single-step process and to produce high-value amino acid supernatant without having an isolated enzyme. In our study, we show the effect of 2.5% (vol/vol) CTLS supernatant obtained from Y. lipolytica IPS21 on the elongation of the root system of selected plants and provide information on the effect of environmental factors on the efficiency of the bioconversion and the migration of chromium.

Project description:BackgroundBoth industrial biotechnology and the use of cellulosic biomass as feedstock for the manufacture of various commercial goods are prominent features of the bioeconomy. In previous work, with the aim of developing a consolidated bioprocess for cellulose bioconversion, we conferred cellulolytic activity of Yarrowia lipolytica, one of the most widely studied "nonconventional" oleaginous yeast species. However, further engineering this strain often leads to the loss of previously introduced heterologous genes due to the presence of multiple LoxP sites when using Cre-recombinase to remove previously employed selection markers.ResultsIn the present study, we first optimized the strategy of expression of multiple cellulases and rescued selection makers to obtain an auxotrophic cellulolytic Y. lipolytica strain. Then we pursued the quest, exemplifying how this cellulolytic Y. lipolytica strain can be used as a CBP platform for the production of target products. Our results reveal that overexpression of SCD1 gene, encoding stearoyl-CoA desaturase, and DGA1, encoding acyl-CoA:diacylglycerol acyltransferase, confers the obese phenotype to the cellulolytic Y. lipolytica. When grown in batch conditions and minimal medium, the resulting strain consumed 12 g/L cellulose and accumulated 14% (dry cell weight) lipids. Further enhancement of lipid production was achieved either by the addition of glucose or by enhancing cellulose consumption using a commercial cellulase cocktail. Regarding the latter option, although the addition of external cellulases is contrary to the concept of CBP, the amount of commercial cocktail used remained 50% lower than that used in a conventional process (i.e., without internalized production of cellulases). The introduction of the LIP2 gene into cellulolytic Y. lipolytica led to the production of a strain capable of producing lipase 2 while growing on cellulose. Remarkably, when the strain was grown on glucose, the expression of six cellulases did not alter the level of lipase production. When grown in batch conditions on cellulose, the engineered strain consumed 16 g/L cellulose and produced 9.0 U/mL lipase over a 96-h period. The lipase yield was 562 U lipase/g cellulose, which represents 60% of that obtained on glucose. Finally, expression of the hydroxylase from Claviceps purpurea (CpFAH12) in cellulolytic Y. lipolytica procured a strain that can produce ricinoleic acid (RA). Using this strain in batch cultures revealed that the consumption of 11 g/L cellulose sustained the production of 2.2 g/L RA in the decane phase, 69% of what was obtained on glucose.ConclusionsIn summary, this study has further demonstrated the potential of cellulolytic Y. lipolytica as a microbial platform for the bioconversion of cellulose into target products. Its ability to be used in consolidated process designs has been exemplified and clues revealing how cellulose consumption can be further enhanced using commercial cellulolytic cocktails are provided.

Dataset Information

Yarrowia lipolytica and Lactobacillus paracasei Solid State Fermentation as a Valuable Biotechnological Tool for the Pork Lard and Okara's Biotransformation.

Publications

<i>Yarrowia lipolytica</i> and <i>Lactobacillus paracasei</i> Solid State Fermentation as a Valuable Biotechnological Tool for the Pork Lard and Okara's Biotransformation.

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