Production of baker's yeast by fermentation

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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 CH 1.91 O 0.56 N 0.23 ) 10, peptone (empiric formula CH 1.795 O 0.5 N 0.2 ) 10, adjust pH to value of 5.8. Dissolve the glucose in 20 cm 3 of redistilled water and sterilize separately at 120°C for 20 minutes. Dissolve other components of the medium in 60 cm 3 of redistilled water, adjust pH to 5.8 and sterilize at 120 ºC for 20 minutes in a 500 ml Erlenmeyer flask. After the sterilization, blend both parts of the medium aseptically (final volume of 80 ml). After inoculation by one eye from inclined agar, which is used for preservation of the strain, the inoculum is cultivated for 15 hours on a rotary shaker at 26 ºC and at stirring of 200 rpm. 3.2 Preparation of the production medium 3.3 The composition of production medium (in g dm -3 ): d-glucose 25, yeast extract 10, peptone 10. Dissolve the extract and peptone in 1220 dm 3 of redistilled water and pour it into the reactor. Dissolve glucose in 500 dm 3 of redistilled water and pour it into 1000 ml Erlenmeyer flask. Check the completeness of the reactor and test-connect it according to scheme (Fig.1.) 8 7 6 out-gassing pO T air 3 2 5 4 1 pH FERMENTOR Fig.1. Scheme of connections of the fermentor 9 10 B I O S T A T 1 – safety vessel with glass wool, 2 – U-manometer, 3,5,7 – safety vessel, 4,8 – sterile air microfilters, 6 – condenser, 9 – storage vessel containing the acid for adjustment of pH, 10 – storage vessel for inoculum. 4
3.4 Fermentation (conditions 28°C, 400 RPM, 1.5 VVM) • All sensors, except for temperature sensor, are sterilizable and would be sterilized right inside the reactor. The pH sensor must be calibrated before the sterilization in a subprogram of the main menu CALIBRATION. • Calibration of the pH sensor: use an arrow on the keyboard and set the cursor to second row and switch the regime from “auto” to “man”. Switching is performed using button ALTER and the choice is confirmed by button ENTER. The cursor relocates to temperature; manually enter desired temperature. Press ENTER. Immerse the pH sensor into pH 7 buffer and press ENTER. When the pH value stabilizes, the biostat would record the value and it will automatically relocate the cursor to following row. Immerse the electrode into pH 4 buffer and press ENTER. When the pH value stabilizes, the regime is automatically switched to “auto” and the calibration is successfully finished. CALIBRATION pH : 7.2 actual pH value inside reactor TEMP: 28 C man Temperature at calibration. - auto: calibration is performed at actual temperature inside reactor - man: temperature is set manually BUFZ: 7.00 pH ok Displaying of pH of “zero” buffer Ok – confirmation of the calculation of zero point. The cursor automatically skips to row BUFS BUFS : 4.00 pH ok Displaying of pH value for calculation of the slope. Ok – after confirmation, the cursor skips to second row and the regime automatically switches to “auto”. Connections of sensors at the head of reactor is displayed on Fig.2. 1 – inlet of air 2 – outlet of air 3 – condenser 4 – access to temperature sensor 5 – access to oxygen sensor 6 – access to pH sensor 7 – sampling port 8 – inlet of acid 9 – inoculum 10 – pouring of mediums Fig.2. Scheme of the head of bioreactor 5
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Page 7 and 8: Calibration of the oxygen sensor: s
Page 9 and 10: 3.6 Measurement of optical density
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Production of baker's yeast by fermentation

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