Tips for pre-treatment of fermentation broths

Fermentation broth pretreatment is an important step in the extraction of biochemical substances from microbial fermentation broth. The aim is not only to separate cells, cells and other suspended particles, but also to remove some soluble impurities and change the properties of the filtrate to facilitate subsequent steps.

The method of pretreatment depends entirely on the properties of the separable substance, such as its stability to PH and heat, whether it is protein or non-protein, the mass and size of the molecule, and so on. The specific methods are mainly as follows:

1. Heating method

The heating method is the simplest and most inexpensive method of pretreatment, in which the suspension is heated to the desired temperature and kept warm for an appropriate period of time. According to the principle of fluid mechanics, the rate at which the filtrate passes through the filter cake is inversely proportional to the viscosity of the liquid, which shows that reducing the viscosity of the liquid can effectively increase the filtration rate.

At the same time, the protein can be coagulated at the appropriate temperature and heating time to form larger particle agglomerates, which further improves the filtration characteristics of the fermentation broth. For example, after adjusting the acid to PH3.0, the viscosity of the streptomycin fermentation broth is heated to 70 °C for half an hour, and the filtration rate can be increased by 10~100 times.

When using the heating method, the heating temperature and time must be strictly controlled. First of all, the heating temperature must be controlled within the range that does not affect the activity of the target product; Secondly, if the temperature is too high or the time is too long, the cells will be dissolved, and the intracellular substances will overflow, which will increase the complexity of the fermentation broth and affect the subsequent separation and purification of the products. Therefore, the key to the heating method depends on the thermal stability of the product.

2. Adjust the PH value of the suspension

ADJUSTING THE PH VALUE OF SUSPENSION IS ONE OF THE MOST COMMONLY USED METHODS FOR FERMENTATION BROTH PRETREATMENT IN THE FERMENTATION INDUSTRY. The pH value directly affects the ionization degree and charge properties of some substances in the fermentation broth, so proper adjustment of the pH value of the fermentation broth can improve its filtration characteristics. For amphoteric substances such as Aa and Pro, the solubility is the smallest at the isoelectric point, which is the isoelectric precipitation method.

For example, in the production of monosodium glutamate, isoelectric point precipitation is used to extract glutamate; In membrane filtration, the macromolecular substances in the fermentation broth are easily adsorbed with the membrane, and the plugging and pollution can be reduced by adjusting the pH value to change the charge properties of the adsorbed molecules.

3. Coagulation and flocculation

Both coagulation and flocculation are important methods for suspension pretreatment. The treatment process is to add the chemical agent to the suspension in advance, change the dispersion state of colloidal particles such as cells, cell debris, bacteria and Pro, destroy their stability, make them coagulate into larger particles, facilitate the improvement of filtration rate, and can effectively remove impurities and solid impurities, and improve the quality of the filtrate. However, coagulation and flocculation are two different methods, and their specific treatment processes are still different, which should be clearly distinguished and not confused.

Coagulation refers to the process of adding a certain electrolyte to the colloidal suspension, and under the action of electrolyte heteroion, the electric double layer potential of the colloidal particles decreases, so that the colloids lose stability and make the particles condense into a lumpy condensate of about 1mm in size.

Flocculation refers to the process of using flocculants to cross-link colloidal particles into a network to form a flocculation group of about 10mm in size. Among them, the flocculant mainly plays the role of bridging. The particles of the condensate obtained by the coagulation method are often only about 1 mm, which is relatively small, and sometimes cannot be effectively separated. The flocculation method can often form a coarse floc (about 10mm), which makes the fermentation broth easier to separate.

4. Add filter aids

Filter aid is an incompressible, porous particle that loosens the filter cake and increases the filtration rate. This is because after the use of the filter aid, a large number of fine particles in the suspension are adsorbed to the surface of the filter aid, which changes the structure of the filter cake, so that the compressibility of the filter cake is reduced, and the filtration resistance is reduced.

Commonly used filter aids are diatomaceous earth, cellulose, asbestos powder, perlite, clay, charcoal particles and starch. Among them, the most commonly used is diatomaceous earth, which has great adsorption and permeability capacity, can filter out 0.1~1.0μm particles, and has stable chemical properties, which is not only an excellent filter medium, but also an excellent filter aid.

5. Add reactants

In some cases, the effect of certain impurities in the fermentation broth on filtration can be eliminated by adding some reactants that do not affect the product of interest, thereby increasing the filtration rate.

The added reactant reacts with certain soluble salts to form insoluble precipitates, such asCaSO4. The precipitate prevents the bacteria from sticking and gives the hyphae a lumpy structure, and the precipitate itself acts as a filter aid and solidifies the colloids and suspended solids, thus improving filtration performance. If the reaction agent and reaction conditions can be selected correctly, the filtration rate can be increased by 3~10 times.

If the fermentation broth contains insoluble polysaccharide substances, it is best to convert it into monosaccharides with enzymes first to increase the filtration rate.

For example, vancomycin uses starch as a medium, 0.025% amylase is added before filtration of the fermentation broth, stirred for 30min, and 2.5% diatomaceous earth is added as a filter aid, which can increase the filtration rate by 5 times.