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Production of baker’s yeast by fermentation

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1. THEORY

The primary task of bioengineering is to design and operate bioreactors with the best possible economic effect. Such optimization is not possible without solving material balances and energetic balances of given bioprocess. At design and process control of production, material balances enable optimization of effectiveness ofsubstrate transformation to products. Thus, the costs of raw materials could be optimized. Similarly, energetic balances help at optimization of stirring costs and of costs of bioreactor cooling. Material and enthalpy balances of bioreactors might not be formulated without so-called source terms, which reflect the influence of biochemical process on balance of components and energy. At classic chemicalreactions, it is relatively straightforward to identify all the reactions and their stoichiometry. However, the principles of classic stoichiometry are rather hard to apply at biotechnological productions. The reason for this is that thousands of reactions take place simultaneously in bioreactors. It is impossible to identify or quantify all of these reactions. Thus, the description ofstoichiometry and of bioprocess kinetics needs to be simplified. This means that we describe only the transformation of key substrates to key products. The key substrates are especially the sources of carbon, nitrogen, oxygen and also the sources of phosphorus and sulfur. Solution of such stoichiometric equation is based on balance of atoms of carbon, nitrogen, oxygen and hydrogen, which might be replaced bybalance of free electrons. Remaining equations, which are necessary for calculation of stoichiometric coefficients, could be obtained from experimental data, such as the respiratory quotient RQ or yield factors Yx/s, Yp/s, YO2/s. For better illustration, an example is introduced: a stoichiometry of aerobic cultivation of onecell proteins from bacteria Methyllococus capsulatus. As a source ofcarbon, methane was used; ammonia was the source of nitrogen and no other product was formed in the liquid phase. The overall process could be described by a simple equation:

s CH4 + a NH3 + b O2 = c CH1.8O0.5N0.2 + d CO2 + e H2O

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Where the values s, a, b, c, d, e are modified stoichiometric coefficients related to 1 mole of consumed substrate Yi/s (mol i/mol s). These are actually theyield factors defined as a ratio

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of reaction rates. In order to solve a stoichiometric equation with 6 unknown stoichiometric coefficients, six equations are necessary. These equations are the balances of atoms of C, N, H, O, s = 1 (modified coefficients related to 1 mole of consumed methane). Another equation is one for calculation of average yield factor Yx/s, which could be calculatedfrom experimental measurement of rate of substrate consumption and of biomass production (Yx/s = 0.4785). Balance of elements’ atoms comprises a system of six algebraic equations, which could be solved. Thus, it is possible to calculate e.g. the amount of oxygen and nitrogen necessary for preparation of given amount of biomass.

C : 1s + 0 + 0 − 1c − 1d + 0 = 0 N : 0 + 1a + 0 − 0.2c + 0 + 0 = 0 O:H: S: 0 + 0 + 2b − 0.5c − 2d − 1e = 0 4s + 3a + 0 − 1.8c + 0 − 2e = 0 1s + 0 + 0 + 0 + 0 + 0 = 1

Yx/s: 0 + 0 + 0 − 1c + 0 + 0 = 0.4785

2. Aim of the work
1. To perform an aerobic cultivation of baker’s yeast in a laboratory stirred batch fermentor. 2. To experimentally monitor development of concentration of substrate, biomass and oxygen during the process. To calculate average yieldfactors based on these measurements. 3. To construct a stoichiometric equation of the process. To calculate the values of modified stoichiometric coefficients and calculate the total amount of consumed oxygen and produced CO2.

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3. METHODS
3.1 Preparation of the inoculum
The composition of inoculation medium (in g dm-3): d-glucose 25, yeast extract (empiric formula CH1.91O0.56N0.23) 10,...