CH640393A5 - METHOD FOR REFINING TOBACCO. - Google Patents

Description

The invention relates to a process for refining tobacco by breaking down nitrates, nitrites and other nitrogen compounds contained in the tobacco.

Many tobacco, e.g. Burley, contain unwanted nitrates and / or nitrites. Fermentation processes are known in which nitrogen compounds are broken down enzymatically, but only to a very small extent and also only as a side effect to other enzymatic reactions.

The object of the invention is to break down nitrogen compounds, in particular nitrates and nitrites, down to an acceptable residual content.

The inventive method is defined in claim 1.

The microorganisms remain in their exponential growth phase under the process conditions, as long as the required nitrogen requirement can be met from the nitrogen compounds, such as nitrates and nitrites.

The microorganisms used are forced to cover their nitrogen requirements from the nitrates, nitrites and other nitrogen compounds to be broken down. The nitrogen that is in the nitrate or nitrite is used by the microorganisms to build up the organic compounds necessary for their life. These are mainly amino acids and proteins that do not interfere with smoking, on the contrary, they can have a beneficial effect on the taste of smoke.

Since the added culture is already in its exponential growth phase, its microorganisms have a lead of about 8 hours over other undesirable microorganisms that are still in their latent phase. Such undesirable microorganisms cannot achieve this advantage within the exposure time, which is limited to a maximum of 24, preferably 16 hours, to such an extent that they can develop a noteworthy effect. In this way it can be ensured that no undesirable side effects occur.

Unfermented, air-dried tobacco often has a nitrate content of around 50 g per kg dry weight. In extreme cases, amounts of nitrate up to 80 g per kg dry weight were measured. The desired residual content depends on the reuse of the tobacco. In the case of tobacco varieties that make up a significant part of the ready-to-smoke tobacco mixture, the desired residual content 55 is at most 5 g of the anions nitrate and nitrite per kg dry weight; for tobacco varieties that make up only a small proportion of the ready-to-smoke mixture, the desired residual content may be somewhat higher. In general, the desired residual content is in the range from 3 to 20%, preferably 60 to 5%, of the original content of the tobacco to be refined, based on the total weight of the anions nitrate and nitrite.

The undesired action of any other microorganisms that may be present due to contamination can be prevented even more by bringing the culture of the microorganisms into action so intensively that the nitrates and nitrites are reduced to a minimal residual content within 6 to 16 hours and that

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the effect of the microorganisms is stopped immediately as soon as the desired residual content is reached. is enough.

The minimum residual content, the residual content that can be achieved with the invention without exceptional measures, depends on the quality of the tobacco to be refined and is generally 0.01 to 0.1% of the original content of the tobacco to be refined, in each case based on the total weight of the anions nitrate and nitrite.

The impact of the microorganism culture can be intensified by optimizing the living conditions for these microorganisms with regard to temperature, humidity, pH environment and nutrient supply and by bringing the culture into action in high concentration on the nitrates and nitrites from the outset. The optimal conditions can be found by trying. The degradation of the nitrates and nitrites to the desired or minimum residual content can be analytically verified.

The action can be stopped by not maintaining the living conditions of the microorganisms, for example by sharply lowering the temperature, drying them, or by removing the microorganisms, for example by filtering them off as active sludge, if the action takes place in a liquid medium.

The microorganisms used can be bacteria, for example those from the genus Aerobacter, Pseudomonas, Micrococcus or Echerichia, but they can also be fungi, for example those from the genus Rhodutorula or from the genus Candida.

Bacteria belonging to the normal micro flora of tobacco leaves are preferred because they have a particularly fast denitrifying effect and do not change the tobacco in an undesirable manner.

A corresponding embodiment of the invention is characterized in that a pure culture of bacteria is used, which is obtained by inoculating an aqueous swab of nitrate-containing leaves or rotten leaves of a nutrient solution which contains the nitrogen content required for the incubation predominantly in the form of nitrates, is buffered to pH 6.6 to 7.5 and is incubated at 25 to 35 ° C for 6 to 16 hours by shaking under sterile intensive aeration and then used as active inoculum for inoculating a fresh such nutrient solution, with which the incubation is then carried out is repeated and so on until the pure culture is formed.

The smear is preferably made from tobacco leaves. But you can also get a usable smear if you use forest soil that has arisen from mouldered leaves or contains mouldered leaves.

A pure culture can be obtained by this process, the microorganisms of which are in an active, ie exponential, growth phase. This can either be used immediately or kept and deactivated for this purpose and then reactivated before further use.

The invention can also be advantageously used in connection with a pure culture of the Enterobacter ae-rogenes bacterium, preferably from the standard strain ATCC 13048. Pure cultures of this standard strain are available from the German Collection of Microorganisms, Göttingen, Grise-Bachstrasse 8, under the DSM image no. DSM 30053 released and available for everyone without restriction. This microorganism strain belongs to the prior art and is not the subject of the invention.

Enterobacter aerogenes have the following properties:

Motile sticks, 0.3-1.5 | im;

Gram - gas evolution at 37 ° C

Glycerin +

Inositol +

Andonitol +

Voges-Proskauer +

Methyl red -

Phenylamine deaminase -

Urease -

Catalase +

Ornithine decarboxylase +

Lysine decarboxylase +

Hydrolysis of aesculin + growth in the presence of KCN + on malonate as the only one

Carbon source +

In natural tobacco, the nitrates and nitrites are difficult to access for external microorganisms, which is why an embodiment of the invention is recommended, which is characterized in that nitrates and nitrites and other water-soluble constituents are dissolved out of the tobacco to be refined with water, so that the tobacco extract solution thus obtained is inoculated with the microorganism culture and mixed with the nutrient solution under sterile intensive ventilation for 6 to 16 hours under favorable aerobic living conditions for these microorganisms and that the action of the microorganisms is then immediately stopped and that the solution components contained in the low-nitrate tobacco extract solution tobacco washed out with water can be added.

In many cases, for example in the reprocessing of tobacco waste, the tobacco is pulverized, slurried and formed into a film with binders, which is then solidified by drying. In such a case, the microorganisms can advantageously be allowed to act while the tobacco is suspended in the suspension, and this is preferably done by grinding the tobacco to be refined and suspending it with water and adding the microorganism culture and nutrient solution with sterile intensive aeration for 6 to 16 Is kept for hours under favorable aerobic living conditions for the microorganisms and then immediately the action of the microorganisms is stopped by shaping the suspension into or on foils and drying to a moisture content of 10 to 30%.

The microorganism culture is preferably used as a pure culture, the degree of purity having to be sufficient to avoid significant side effects. The active sludge obtained in microbiological degradation is preferably added to tobacco and inactivated. This can be done, for example, by spraying the active sludge on tobacco leaves and drying it.

The invention will now be explained in more detail with the aid of a few examples.

example 1

Obtaining the pure culture in the culture process

20 g (grams) of D-glucose; 6.4 g NaCl; 3.5 g KN03; 4.5 g KH2PO4 and 23.5 g Na2HPÓ4 • 2H20 are dissolved in 11 (liters) water. The nutrient solution obtained in this way is divided into 5 equal portions I to V of 200 ml (milliliters) each,

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each portion is placed in a 500 ml Erlenmeyer flask which is closed with a porous stopper. Then the whole is sterilized and stored at 20 ° C.

100 g dry Burley tobacco leaves are washed with 500 ml water under sterile conditions. 1 ml of the washing suspension formed is drawn off sterile and added to portion I of the nutrient solution. This portion I is incubated on a shaker for 16 hours at 30 ° C under sterile conditions with an air flow rate of 200 ml • min-1 (milliliters per minute). It is then enriched by taking 1 ml of this portion under sterile conditions and inoculating into portion II of the nutrient solution. Portion II is then treated like Portion I and, after incubation, 1 ml of Portion II is inoculated into Portion III and this portion is treated accordingly and so on up to and including Portion V.

After the portion V has been incubated for 16 hours as described, it contains the pure culture of the microorganisms of an Enterobacter strain which covers its nitrogen requirement by nitrogen obtained by the degradation of nitrogen compounds. It mainly breaks down nitrates, nitrites and some nitrosamines. Is not that the case,

then everything is discarded and the extraction repeated from the beginning with other Burley tobacco leaves. The microorganisms of this pure culture are in their exponential growth phase at this time and up to 8 hours later.

Refinement of tobacco ribs

1 kg of Maryland tobacco, in which the nitrates and nitrites are to be reduced, is stripped. That makes 250 g ribs and 750 g strips.

The 250 g ribs are washed with 1250 ml of 70 ° C warm water. Water-soluble nitrogen compounds present in the fins are extracted together with other water-soluble components. The rib extract solution thus obtained is drawn off from the ribs, placed in a 21-Erlenmeyer flask which is closed with a porous stopper, cooled to 30 ° C., mixed with 12.5 g of D-glucose and with 10 ml of the above-mentioned Pure culture inoculated within the period in which the microorganisms of this pure culture are still in their exponential growth phase.

The inoculated rib extract solution is aerated at 30 ° C on a shaker under sterile conditions with an air throughput of 1300 ml • min-1 and thereby incubated for 16 hours until the anions nitrate and nitrite are broken down to a content of 0.1 g per 1 are. Other nitrogen compounds are also broken down in the process.

The resulting low-nitrate rib extract solution is then immediately centrifuged, whereby the microorganisms settle, which are then removed as active sludge.

The centrifugate from the low-nitrate rib extract solution is added to the pre-dried, washed-out ribs, which are then dried to 20% moisture content. Since the centrifugate still contains the remaining tobacco constituents detached from the fins, these reach the fins again, so that the fins then essentially again contain their original constituents, with the exception of the degraded nitrogen compounds. In this way, ribs rich in nitrate and nitrite are converted into ribs low in nitrate and nitrite.

Example 2

As example 1 with the only difference that the inoculated rib extract solution is incubated for 20 hours until 50% of the nicotine originally contained in the rib extract solution has been broken down. The rib extract solution then contains at most traces of nitrate and nitrite because the microorganisms used only start to break down nicotine when there is no longer any nitrite or nitrate available.

Example 3

Like example 1, with the only difference that the centrifugate of the low-nitrite rib extract solution is not added back to the ribs from which the extract was obtained, but to other ribs which are treated in the same way.

Example 4 Refinement of Tobacco Leaves 1 kg of Maryland tobacco, in which the nitrates and nitrites are to be reduced, is stripped. That makes 250 g ribs and 750 g strips. The 250 g ribs are treated as indicated in Example 1. The 750 g strips are individually immersed in 1250 ml of 50 ° C. warm water for 20 seconds. This removes the water-soluble compounds present in the superficial areas of the strips. Since the nitrates and nitrites contained in the strips are mainly contained in the superficial areas, they largely get into the water. In the strip extract solution obtained in this way, which also contains other nitrogen compounds, the nitrogen compounds are degraded microbiologically until the anions nitrate and nitrite are broken down to a content of 0.1 g per liter.

The strip extract solution is then concentrated and the strips are added again by spraying. For this purpose, the strip extract solution is treated like the rib extract solution according to Example 1.

Example 5

Obtaining the pure culture as in Example 1 Refining tobacco leaves 250 g of Burley tobacco leaves are washed with 1250 ml of 50 ° C. warm water. The resulting tobacco extract solution is further treated like the rib extract solution according to Example 1, and the low-nitrate tobacco extract solution finally obtained is centrifuged off from the active sludge and added to the tobacco leaves again.

Example 6

1 kg of tobacco waste, in which the nitrates and nitrites are to be reduced, is finely ground to a maximum grain size of 150 [i. 150 ml of active sludge, which are obtained according to Example 1 and are still in their exponential growth phase, are kept in this phase by the active sludge immediately together with 30 g of D-glucose, 9.6 g of NaCl, 675 g of KH2PC> 4 and 35, 25 g Na2P04 • 2H20 is added to 1500 ml water. This gives a suspension of active microorganisms into which the tobacco powder is then stirred. The resulting slurry is aerated for 24 hours at 30 ° C under sterile conditions with an air pressure set of 3000 ml • min -1 in a 101 Erlenmeyer flask which is closed with a porous stopper. The microorganisms in the tobacco powder break down nitrates and nitrites and other nitrogenous compounds down to 1/10 of the original content. Immediately afterwards, 150 g of carboxymethyl cellulose are introduced into the slurry, and the slurry is spread into a 3 mm thick layer and dried to 15% moisture content. This stops the action of the microorganisms and turns the slurry into foils

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regenerated tobacco solidified, which can be processed. The regenerated tobacco obtained is low in nitrates and nitrites.

Example 7

Like example 6 with the only difference that, as in example 2, the breakdown is down to a nicotine content of 50%.

Example 8

Like example 1 with the only difference that the inoculated rib extract solution is incubated at 30 ° C on a shaker instead of 16 hours for only 8 hours, so that the anions nitrite and nitrate are degraded to a lesser extent than according to example 1.

The obtained low-nitrate rib extract solution is then further treated as in Example 1 and added to the ribs which have been pre-dried and washed out as in Example 1 and which are then dried to a moisture content of 20%. The ribs treated in this way contain a third of the original content of nitrate and nitrite.

Example 9

Refinement of tobacco ribs as in Example 1 with the only difference that the inoculation takes place with a pure culture of the bacterium Enterobacter aerogenes ATTCC 13048.

Example 10

As in Example 9, with the only difference that the centrifugate of the low-nitrite rib extract solution is not added back to those ribs from which the extract s was obtained, but to other ribs which were treated in the same way.

Example 11

Refinement of tobacco leaves as in Example 4 with the only difference that the inoculation is carried out with a pure culture of the bacterium Enterobacter aerogenes ATTCC 13048.

Example 12

15 Refinement of tobacco leaves as in Example 5 with the only difference that the inoculation is carried out with a pure culture of the bacterium Enterobacter aerogenes ATTCC 13048.

20 Example 13

As example 6 with the only difference that active sludge, which was obtained according to example 9, is used.

25 Example 14

As example 8 with the only difference that the inoculation is carried out with a pure culture of the bacterium Enterobacter aerogenes ATTCC 13048.

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Claims (7)

640393 2nd PATENT CLAIMS 1. A process for refining tobacco by breaking down nitrogen-containing compounds contained in the tobacco, characterized in that a culture of microorganisms which need nitrogen and can cover their nitrogen requirement by breaking down nitrogen-containing compounds are shifted to their exponential growth phase under aerobic living conditions Nutrient solution for this culture, which has no essential nitrogen source accessible to the microorganisms of this culture, is brought into contact with the tobacco components under favorable aerobic living conditions for the microorganisms of this culture until the nitrogenous compounds have broken down to the desired residual content, and then immediately , after 24 hours at the latest, the effects of the microorganisms are stopped. 2. The method according to claim 1, characterized in that the nitrogen-containing compounds comprise nitrates and nitrites. 3. The method according to claim 2, characterized in that the culture of the microorganisms is so intensely affected that the nitrates and nitrites are broken down to a minimal residual content within 6 to 16 hours and that the action of the microorganisms caused is stopped immediately as soon as the desired residual salary can be reached. 4. The method according to any one of claims 1 to 3, characterized in that a pure culture of bacteria is used, which is obtained by inoculating an aqueous swab of nitrate-containing leaves or decayed leaves of a nutrient solution which contains the nitrogen content required for the incubation predominantly in Contains form of nitrates, is buffered to pH 6.6 to pH 7.5 and is incubated at 25 to 35 ° C for 6 to 16 hours by shaking under sterile intensive aeration and is then used as an active inoculum to inoculate a fresh such nutrient solution , with which the incubation is then repeated and so on until the pure culture is formed. 5. The method according to any one of claims 1 to 3, characterized in that a pure culture of the Enterobacter aerogenes bacterium from the ATCC 13048 strain is used. io 6. The method according to any one of claims 2 to 5, characterized in that nitrates and nitrites and other water-soluble components are dissolved with water from the tobacco to be refined, that the tobacco extract solution thus obtained is inoculated in isolation with the microorganism culture i5 and mixed with the nutrient solution underneath sterile intensive aeration is held for 6 to 16 hours under favorable aerobic living conditions for these microorganisms and that the action of the microorganisms is then immediately stopped and that the solution components contained in the low-ni-low-tobacco extract solution are added to water-washed tobacco. 7. The method according to any one of claims 2 to 5, characterized in that the tobacco to be refined 25 oil and slurried with water and mixed with the microorganism culture and nutrient solution under sterile intensive aeration for 6 to 16 hours under favorable aerobic living conditions for the microorganisms and that the action of the microorganisms is then immediately stopped by the suspension to or is formed on foils and dried to a moisture content of 10 to 30%. 8. The method according to claim 6, characterized in that the active sludge obtained tobacco 35 sets and is deactivated.