CS270332B1 - Process for producing yeast products from methanol - Google Patents

Abstract

The session discusses the production of yeast biomass from methanol, which is rich in proteins, essential amino acids and vitamins, and which serves as a valuable protein and vitamin feed for livestock and as a source for the preparation of Cl-specific enzymes such as alcohol oxidase, formaldehyde and formate dehydrogenase, dihydroxyacetone synthase, as well as catalase, NADH, GSII and ATP from methanol. "The method of producing yeast products allows to increase the yield of yeast biomass from methanol, to increase the concentration of cells in the medium and to shorten the cultivation time. In the production of yeast products from methanol, methylotrophic yeast strains are used, in particular Candida boidinii 2 and Hansenula ep.3, while the cultivation medium contains, in addition to methanol in an amount of 0.5 to 1% by weight, molasses distillery stills in an amount of 0.2 to 0.3% by weight. Cultivation takes place at a temperature of 30 to 40 °C and a pH of 3.0 to 3.5.

Description

The invention relates to a method for producing yeast products from methanol.

Molasses distillery wastes are commonly used to produce feed yeast. They are a valuable raw material because, in addition to being a source of carbon, they contain a large amount of potassium, assimilable nitrogen and sometimes a sufficient amount of magnesium. Their content of growth substances, which come into the wastes from beet molasses, but also from yeast, during fermentation in the distillery, and the presence of a number of trace elements are important. The disadvantage of molasses distillery wastes as a raw material for the production of feed yeast proteins is their unbalanced composition, low carbon content and the need to add a large amount of acids to adjust the pH of the fermentation medium during cultivation. Therefore, molasses distillery wastes are preferably used as an addition to the medium in the production of yeast proteins from sulphite leachates, from ethanol-sulphite mixtures and also from synthetic ethanol using methylotrophic strains other than yeast. Molasses distillery stills serve here as a stimulator of yeast growth, as a source of potassium and possibly as a neutralizing agent (see author's certificate No. 203 413 and No. 174 431). The subject of the invention is a method for producing yeast products from methanol, the essence of which is that methylotrophic yeast strains are cultivated in a medium containing molasses distillery stills in an amount of 0.2 to 0.3% by weight and methanol in an amount of 0.5 to 1% by weight, at a temperature of 30 to 40 °C and a pH of 3.0 to 3.5.

As methylotrophic yeast strains, Candida boidinii llBh, C. boidinii 1, Hansenula polymorpha, Pichia or mixtures thereof under sterile culture conditions and/or acid-tolerant methylotrophic yeast strains Candida boidinii 2 and Hansenula sp. 3 under non-sterile culture conditions can be advantageously used.

Molasses distillates serve not only as a source of growth substances and mineral salts for the growth of methylotrophic yeasts on methanol media, but also shorten the time of yeast cultivation. Methylotrophic yeasts in the presence of molasses distillates grow on methanol media immediately after ionoculation at a maximum growth rate. When inoculating the inoculum onto a new methanol medium without molasses distillates, methylotrophic yeasts show a 2 to 6 h lag, during which they almost do not grow, depending on the size of the inoculum and aeration. Furthermore, molasses distillates increase the yield of yeast biomass above 40%, which is the maximum yield of methylotrophic yeasts, based on the consumed methanol. In the presence of molasses distillates, it is possible to increase not only the yield of biomass, but also the yield of all C^ specific products that are produced from the obtained yeast biomass. When enriching the medium with atlas stillage, in conventionally stirred and aerated fermenters, using the inflow method of dosing small amounts of methanol into the fermenter, it is possible to achieve a yeast cell concentration of over 40 g of dry biomass in 1 liter of fermentation liquid, i.e., 10 g in 1 liter of medium more than in a medium with methanol alone. By using molasses stillage in methanol fermentations, it is possible, as in the case of sulfite or alcohol mixtures, to replace part of the potassium ions added to the medium in the form of potassium phosphate or potassium hydroxide with the potassium content in the stillage, thus saving these raw materials.

Biomass obtained on a medium with methanol and molasses distillates is identical in terms of nutritional value and protein content to biomass from methanol alone, i.e. it contains 60 to 62% crude protein rich in essential amino acids such as lysine, leucine, threonine, valine, and vitamins B^ B ₂ and also contains the maximum level of specific enzymes such as alcohol oxidase, formaldehyde and formalin dehydrogenase, dihydroxyacetone synthase, catalase, reduced nicotinamide adenine dinucleotide (NADH) and reduced glutathione (GSH).

In principle, there are two ways to proceed with cultivation. Molasses distillates can be added to the mineral medium either once at the beginning of fermentation, or gradually during cultivation in a concentration of 0.1 to 0.3% by weight.

CS 270332 Bl

Example 1

In a laboratory mechanically stirred fermenter with a total volume of 5 l and a filling of 2.5 l, a non-sterile single-time cultivation of the yeast Candida boidinii 2 on a mineral medium with methanol was carried out. The composition of the mineral medium was: 1 liter of tap water, 3 g NH4CI; 7 g _KH2PO4 ; 0.5 g _MgSO4.7H2O ; 0.1 g NaCl; 0.1 g yeast extract and 10 μg biotin. At the beginning of the cultivation, methanol was added to the culture medium in an amount of 1% by weight, and molasses distillate in an amount of 0.25% by weight. During the cultivation, the pH of the medium was maintained at a constant level of 3.0 by automatic dosing of a 10% NaOH solution and the temperature was maintained at 30 °C. The aeration of the fermenter was 0.5 to 1.0 liters of air per hour, the number of revolutions of the stirrer was 600 per second.

The concentrated molasses stillage contained potassium at a concentration of 11% by weight and total nitrogen at a concentration of 5% by weight. The concentration of yeast cells after inoculation of the medium in the fermenter was 0.2 g per liter.

The addition of molasses cake to the methanol medium completely eliminated lag, increased the growth rate of cells in the initial stages of growth, and increased the biomass yield based on methanol from 38% to 45% by weight. On the molasses cake medium without methanol, C. boidinii 2 yeast cells grow with a 4% biomass yield.

For comparison, three more cultivations were carried out with the amount of molasses jyýpalk, as shown in Table 1.

Table 1 , i'f· ' -

Effect of molasses distillates on the biomass yield of the yeast C. boidinii 2 from methanol.

Molasses shots -1 dry weight (g.l 0 1.0 2.5 3.0 Increase in dry cell mass (group ¹ ) 3.8 ' 4.2 4.5 4.6 biomass yield from methanol (%) 38 42 45 46

Example 2

In a 75 l volume, mechanically stirred fermenter filled with 30 to 50 l of mineral medium of the composition as in Example 1, a non-sterile single-use cultivation of the yeast Candida boidinii 2 was carried out. Methanol was dosed via an inflow. method in the form of a methanol-lye mixture (methanol and NaO), preferably a methanol-ammonia mixture (methanol and ammonia water) depending on the pH of the medium, which was maintained automatically at a constant level of 3.0 to 3.2. Molasses pellets were dosed separately and gradually during the cultivation in an amount of 2.5 g per liter of medium once every 24 hours. The air flow and the speed of the stirrer were regulated so that the concentration of dissolved oxygen in the medium was in the range of 10 to 60% saturation. The cultivation temperature was as in example 1, i.e. 30 °C. The amount of biomass obtained in the fermenter was 40 to 45 g of dry cell mass in 1 liter of medium.

Example 3.

The procedure was the same as in example no. 2, with the difference that the yeast strain Hansenula sp. 3 was used at a cultivation temperature of 37 to 40 °C. :

CS 270332 Bl

Example 4

In a 75 l, mechanically stirred fermenter with a volume of 30 to 50 l of mineral medium of the composition as given in Example δ. 1, non-sterile continuous cultivation of the yeast Candid* boidinii 2 was carried out. The growth rate of the medium was in the range of D 0.05 h” ¹ to 0.15 h” ^3, preferably at D 0.1 h ¹ . Methanol in a concentration of 0.5% to 1% by weight, and molasses distillates in a concentration of 0.25% by weight, were dosed into the fermenter together with the medium. The pH of the medium and the cultivation temperature were maintained automatically as in Example No. 1. The yield of yeast biomass was 42 to 45%.

Example 5 ·

The procedure was the same as in Example No. 3, with the difference that methanol was dosed separately from the medium at a concentration of 25% by weight, depending on the dilution rate and the concentration of dissolved oxygen in the medium, which was maintained automatically in the range of 20 to 40% saturation, and the pH of the medium was maintained at a constant pH of 3.0 to 3.2 by automatic dosing of 50% NaOH, KOH or ammonia water.

Example 6

The procedure was the same as in Example No. 5, with the difference that the yeast strain Hansenula sp. 3 was used at a cultivation temperature of 37 to 40 °C.

The yeast biomass obtained by the method according to the invention is rich in proteins, essential amino acids and vitamins and can be advantageously used as a valuable protein and vitamin feed for livestock · as a source for the preparation of specific enzymes such as alcohol oxidase, formaldehyde-a formate dehydrogenase, dihydroxyacetone synthase, as well as catalase, nicotine adenine dinucleotide, adenosine triphosphate (ATP) and NASH, GSH, from methanol using a single-shot, feed-through and continuous method of cultivating methylotrophic yeasts. <

Claims (3)

1. A method for producing yeast products from methanol, characterized in that the methylotrophic yeast strains are cultivated in a medium containing molasses distillery stills in an amount of 0.2 to 0.3% by weight and methanol in an amount of 0.5 to 1% by weight, at a temperature of 30 to 40 °C and a pH of 3.0 to 3.5. .2. The method according to item 1, characterized in that the methylotrophic yeast strains used are Candida boidinii llBh, C. boidinii 1, Hansenula polymorpha, Pichia or mixtures thereof under sterile conditions and/or acid-tolerant methylotrophic yeast strains Candida boidinii 2 and Hansenula sp. 3 under non-sterile culture conditions.