The development a medium for the cultivation of lactic acid bacteria using sweet potatoes

Lactic acid bacteria (LAB) are the most important microorganisms typically associated with the human gastrointestinal tract and used in animal feeds and human foods. They play the main role in food fermentation processes. They are generally recognized as safe (GRAS) and well known as beneficial microorganisms. The fast growing characteristics of LAB is associated with the ability to produce many beneficial compounds such as organic acids and antimicrobial compounds, unique enzymes, and functional compounds [1]. LAB are typically fastidious and require a variety of amino acids, B vitamins, purine and pyrimidine bases for growth.

LAB can be grown and enumerated on agar plates as long as the agar plates are incubated in an oxygen poor environment. Most LAB grow on MRS or M17 (+lactose) agar and form snow white colonies on these media The nutritional requirements of LAB need to be addressed for successful research and industrial application. In addition, consideration must be given the variations in the nutritional requirements of LAB that can occur among species and even strains in the same specie which are also of great concern in LAB research and applications.

Different media are available for LAB; however, the existing media are usually high in price and require several technical steps to achieve high cell density and desirable functionality. The most common standard laboratory media for LAB are de Man-Rogosa-Sharpe (MRS) and M17. MRS was developed to support the growth of Lactobacillus and M17 was developed to support Streptococcus growth. MRS and M17 are known for showing constant support for LAB growth. Such media, MRS and M17, contain complex nitrogen supplementations that are documented to be convenient for LAB growth. However, the use of MRS and M17 is mainly limited to academic purposes owing to their high cost. Therefore, alternative media with lower cost and higher cell density holds high interest for research and industrial purposes [2].

Sweet potatoes are rich with many nutrients and several useful components [3] that could support the growth of LAB. Starch is the main ingredient in sweet potatoes constituting 65±70% of dry weight. In addition, these starchy roots are excellent source of vitamins such as A and C, minerals such as iron and potassium, and fiber [4]. Sweet potatoes (Ipomoea batatas (L.) Lam.) (Batatas an Arawak name) are an abundant agricultural product that is rich in many nutrients. Sweet potatoes are a rich source of carbohydrates (mainly starch and sugars), some amino acids, vitamins (vitamin A, vitamin C, thiamin (B1), riboflavin (B2), niacin, and vitamin E), minerals (calcium, iron, magnesium, phosphorus, potassium, sodium, and zinc), and dietary fiber.

For each experimental replication, 900 g fresh sweet potatoes were used to form 2 L of sweet potato media (SPM). Fresh sweet potatoes were baked in a conventional oven at 400ºC for 1 h. Sweet potatoes were then peeled and blended in a kitchen blinder with deionized distilled water (DDW) at 2:1 ratio (DDW (mL) to sweet potato 100 (g)). This solution was centrifuged at 7800×g for 10 min using Thermo Centrifuge and the supernatant was collected to form sweet potato extract (SPE). SPE was analyzed to confirm nutritional content.

One liter of SPE was mixed with ingredients in Table 1 to form 3 different SPMs (SPM1, SPM2, and SPM3). MRS broth was prepared by dissolving 55 g of MRS broth and 1 g L-cysteine in 1 L of DDW. The addition of L-cysteine was suggested to increase the growth and biomass production of Lactobacillus.

SPMs and MRS were autoclaved Growth of LAB strains in SPM. Since nitrogen sources are main contributors to cost , SPMs were formed at different concentrations of nitrogen sources to lower the cost (see Table 1).

During the first 10 h of incubation, no visual growth was observed. During the next 10 h of incubation, LAB strains showed similar growth rates in MRS, SPM2, and SPM3.

Table 1 - Composition of SPM for 1L of Sweet Potato Extract

#

Component

Components g/L

SPM1

SPM2

SPM3

1

Proteose peptone #3

0

4,0

8,0

2

Beef extract

0

4,0

8,0

3

Yeast extract

0

4,0

8,0

4

Sodium acetate (CH3COONa)

5,0

5,0

5,0

5

Tween 80

1.0 mL

1.0 mL

1.0 mL

6

Disodium phosphate (Na2HPO4)

2,0

2,0

2,0

7

Ammonium citrate (NH4C6H5O7)

2,0

2,0

2,0

8

Potassium phosphate K2HPO4

2,0

2,0

2,0

9

Magnesium sulfate (MgSO4.7H2O)

0,1

0,1

0,1

`10

Manganese sulfate (MnSO4.5H2O)

0,05

0,05

0,05

11

L-Cysteine

1,0

1,0

1,0

SPM1 showed slower growth rates for all tested strains compared to that in MRS. After 20 h of incubation optical density reached averages of 1.505±0.216, 0.984±0.114, 1.427±0.15, and 1.434±0.182 for MRS, SPM1, SPM2, and SPM3, respectively. Results of μmax showed no significant (p > 0.05) differences among MRS, SPM2, and SPM3 for all strains (see Table 15), μmax values in SPM1 were significantly (p < 0.05) lower than that in other media.

Our results revealed that SPM2 could be a suitable medium for the growth of LAB. In addition, SPM2 could be an alternative medium for costly MRS broth. The growth of tested LAB strains in SPM2 and SPM3 was at the same as that in MRS. Since no significant differences were shown between SPM2 and SPM3 with regard to the growth of LAB strains, the formula in SPM2 could be used to form the SPM. These findings may lead to more interest in using sweet potatoes for applications such as fermentation and lactic acid production.

The use of sweet potatoes to develop a new medium for LAB could also open the door to new applications for the use of sweet potatoes.

References:

  1. Von Wright, A., & Axelsson, L. (2011). Lactic acid bacteria: An introduction. In S. Lahtinne, S. Salminen, A. Von Wright & A. Ouwehand (Eds.), Lactic acid bacteria: microbiological and functional aspects (4 ed., pp. 1-17). London: CRC Press.
  2. Aguirre, L., Garro, M. S., & Savoy de Giori, G. (2008). Enzymatic hydrolysis of soybean protein using lactic acid bacteria. Food Chemistry, 111(4), 976-982.
  3. Karna, P., Gundala, S. R., Gupta, M. V., Shamsi, S. A., Pace, R. D., Yates, C., Aneja, R. (2011). Polyphenol-rich sweet potato greens extract inhibits proliferation and induces apoptosis in prostate cancer cells in vitro and in vivo. Carcinogenesis, 32(12), 1872-1880.
  4. Padmaja, G. (2009). Uses and nutritional data of sweetpotato. In G. Loebenstein & G. Thottappilly (Eds.), The sweetpotato (1 ed., pp. 189-234). Belgium, Germany: Springer.
Year: 2018
City: Shymkent
Category: Medicine