Improvement of a Chemically Defined Medium for the Sustained Growth of Lactobacillus plantarum: Nutritional Requirements

Abstract

The aims of this work were to improve a basal synthetic medium (BM) for the growth of Lactobacillus plantarum strains and to establish their amino-acid requirements. Amino-acid use was analyzed in the most nutritionally demanding bacterium. First, the improved BM (L. plantarum synthetic medium [LPSM]) was created by increasing some vitamins in the BM, especially p-aminobenzoic acid, vitamin B₁₂, and biotin; 5-fold phenylalanine, histidine, isoleucine, leucine, lysine, methionine, proline, serine, threonine, and tryptophan; and 10-, 60-, and 75-fold valine, arginine, and tyrosine, respectively. With these additions, the N8 and N4 strains of L. plantarum grew rapidly to reach final cell densities similar to those obtained in Mann–Rogosa–Sharpe medium. When cysteine, leucine, valine, isoleucine, threonine, and glutamic acid were individually removed from this medium, bacterial growth significantly decreased or ceased, indicating that these amino acids are essential for growth. The N4 strain also required lysine and tryptophan in addition to the six amino acids necessary for growth. L. plantarum N4 mainly consumed essential amino acids, such as valine, lysine, cysteine, and threonine as well as the stimulatory amino acid, arginine. Thus, the BM was improved mainly on the basis of annulling limitations with respect to amino acids. With this, improved medium cell densities in the order of 10⁹ colony-forming units/mL have been achieved, indicating that LPSM medium could be used for conducting metabolic and genetic studies on L. plantarum. Their low levels in orange juice suggest that these amino acids may not satisfy the total nitrogen requirement for the development of L. plantarum in the natural environment.