DE2901310A1 - PROCESS FOR REFINING TOBACCO - Google Patents

Description

The invention relates to a method for refining tobacco by breaking down nitrates contained in the tobacco and Nitrites.

Many tobaccos, such as Burley, contain undesirable nitrates and / or nitrites. Fermentation processes are known in the case of which nitrogen compounds are enzymatically processed, but only to a very small extent and only as Side effect to other enzymatic conversions.

It is the task of the inventors to break down nitrates and nitrites down to a beneficial residual content.

The invention is characterized in that a culture of microorganisms that need nitrogen and their Cover nitrogen demand through nitrate and nitrite breakdown can be put into their exponential growth phase under aerobic living conditions with a nutrient solution for them Culture that has no essential nitrogen source accessible to the microorganisms of this culture, under favorable aerobic living conditions for the microorganisms of this culture on the nitrates, nitrites and those accompanying them further tobacco ingredients are brought into action until the nitrates, and other nitrogenous compounds on the desired residual content have been reduced and that then immediately, at the latest after 24 hours, the induced effect of the Microorganisms is canceled. Under these conditions, the microorganisms remain in their exponential growth phase as long as the necessary nitrogen requirements are not met the nitrates and nitrites can be covered.

The applied microorganisms are forced to take their nitrogen requirements from the nitrates to be broken down and cover nitrites. The nitrogen, which is in the nitrate or nitrite, is used by the microorganisms in order to produce them for them

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Build the organic connections necessary for life. Included It is mainly about amino acids and proteins that do not interfere with smoking, on the contrary, one for. the-smoke taste favorable effect. can have.··

Since the added culture is already in its exponential growth phase, its microorganisms have one About 8 hours ahead of other undesirable microorganisms that are still in their latent phase are located. Such undesirable microorganisms can have this advantage within the to a maximum of 24 preferably 16 hours of limited exposure time do not catch up so far that they can develop a noticeable effect. In this way it can be ensured that no undesirable side effects occur.

Unfermented, air-dried tobacco often have a nitrate content of around 50 g per kg dry weight. In extreme cases, nitrate amounts of up to 80 g per kg dry weight have been measured. The desired residual content depends on the further use of the tobacco. In the case of tobacco types that make up a substantial part of the ready-to-smoke tobacco mixture, the desired residual content is a maximum of 5 g of the anions nitrate and nitrite per kg dry weight; in the case of tobacco types that only make up a small proportion of the ready-to-smoke mixtures, the desired residual content can be slightly higher. In general, the desired residual content is in the range from 3 to 20 %, preferably 5 %, of the original content of the tobacco to be refined, based on the total weight of the anions nitrate and nitrite.

The undesirable effects of other microorganisms that may be present due to contamination can be prevented even more strongly by the culture _

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the microorganisms is brought to the effect so intensely that the nitrates and nitrites within 6 to 16 hours are broken down to a minimum residual content and that the induced action of the microorganisms is immediate is canceled as soon as the target residual content has been reached.

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

One can intensify the influence of the microorganism culture by changing the living conditions for it Microorganisms in terms of temperature, humidity, pH environment and nutrient supply are optimized and by brings the culture to action from the outset in a high concentration on the nitrates and nitrites. The optimal Conditions can be found by trying. From the breakdown of nitrates and nitrites to the desired, or the minimum residual content can be verified analytically.

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

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The microorganisms used can be bacteria, for example those from the genus Aerobacter ₉ Pseudomonas, Micrococcus or Echerichia, but also fungi, for example those from the genus Rhodutorula or from the genus Candida.

Preference is given to bacteria belonging to the normal microflora of tobacco leaves because they have a particularly rapid denitrifying effect and do not cause the tobacco to become undesirable Change way.

A corresponding development of the invention is characterized in that a pure culture of bacteria is used which is obtained by taking a, aqueous smear of Nitrate-containing leaves or rotten leaves are inoculated with a nutrient solution that is necessary for incubation required nitrogen content mainly in the form of nitrates, is buffered to pH 6.6 to pH 7.5 and at 25 to 35 ° C (degrees Celsius) for 6 to 16 hours is incubated by shaking under sterile intensive ventilation and then used as an active inoculum for inoculating a fresh such nutrient solution is used, Mt the. then the incubation is repeated and so on until the pure culture is formed.

The smear is preferably made from tobacco leaves. But you also get a usable smear if you go out Forest soil that has rotted leaves or that contains rotted leaves is used «

A pure culture is obtained according to this process, the microorganisms of which are active, i.e. exponential, Are in the growth phase. This is either used immediately or stored and deactivated for this purpose and then reactivated before further use.

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

, Enterobacter aerogenes have the following properties:

Motile double crochet, 0.3-1

Gas evolution at 37 0 _c

Glycerine +

Inositol +

Andonitol + '

Voges-Proskauer + "

Methyl red -

Phenylamine deaminase -

Urease -

Catalase +

Ornithine decarboxylase +

Lysine decarboxylase +

Hydrolysis of esculin + growth ^;

.- in the presence of

KCN - +

on malonate as the only carbon source +

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With natural tobacco, the nitrates and nitrites are for the outside Acting microorganisms are difficult to access, which is why a further development of the invention is recommended, which thereby is characterized in that nitrates and nitrites and other water-soluble components with water from the tobacco to be refined be dissolved out that the tobacco extract solution obtained in this way isolated inoculated with the microorganism culture and with the Nutrient solution is added under sterile intensive ventilation for 6 to 16 hours under favorable aerobic living conditions for this Microorganisms is held and that then immediately the action of the microorganisms is stopped and that the In the low-nitrate tobacco extract solution components contained in the solution are added to tobacco washed out with water.

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

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

The invention is now based on a few exemplary embodiments explained in more detail.

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Example 1 '' · ■

Obtaining the pure culture in the culture process

20 g (grams) D-glucose;

6.4 g NaClj 3.5 g KNO ₃ ; 4.5 _{g of KH 2} PO ₄ and 23.5 g of Na ₃ HPO ₄ . 2Η _£ Ο

are dissolved in 1 l (liter) of water. The nutrient solution obtained in this way is divided into 5 equal portions I to V each 200 ml (milliliters) divided, each portion is placed in a 500 ml Erlenmeyer flask with a porous stopper is closed. Then the whole

sterilized and stored at 20 ° C.

100 g of dry Burley tobacco leaves are mixed with 500 ml of water washed under sterile conditions. 1 ml of the resulting washing suspension is withdrawn in a sterile manner and portion I added to the nutrient solution. This serving I is on a. Shaker 16 hours at 30 C under sterile conditions ventilated with an air flow rate of 200 ml., min. (Milliliters per minute). It is then enriched by taking 1 ml of this portion under sterile conditions and adding it to the Portion II of the nutrient solution is inoculated. The portion II is then treated like the portion I and from the portion JEI- ■ after incubation, 1 ml is inoculated into cue 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 a Enterobacterstammes, which its nitrogen needs by at Removal of nitrogen compounds obtained covers nitrogen. It mainly uses nitrates, nitrites and some Nitrosamines broken down. If this is not the case, then everything will work out discarded and the extraction from the beginning Repeated with other Burley tobacco leaf em. The microorganisms of this pure culture are in their exponential growth phase at this point and up to 8 hours afterwards.

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_ 1 O -

Refinement of tobacco stems

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

The 250 g ribs are mixed with 1250 ml 70 ° C warm water washed. Water-soluble nitrogen compounds that are present in the ribs are thereby combined with other water-soluble ones The rib extract solution obtained in this way is drawn off from the ribs into a

2 1 Erlenmeyer flasks with a porous stopper is sealed, given, cooled to 30 C, with 12.5 g of D-glucose added and with 10 ml of the above-mentioned pure culture, inoculated, within the period of time in which the microorganisms of this pure culture are still in their ^ expqnential "Growth phase.

The inoculated rib extract solution is aerated at 30 ° C on a shaker under sterile conditions with an air flow rate of 1300 ml. Min and thereby incubated, namely 16 hours, until the anions nitrate and nitrite are broken down to a content of 0.1 g per liter . We will also; degraded other nitrogen compounds.

The low-nitrate rib extract solution obtained in this way is then centrifuged immediately, whereby the microorganisms settle, which are then drawn off as active sludge.

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

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Example 2

As in 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 that have set up only begin to break down nicotine when there is no more nirtitol or nitrate available.

Example 3

Like example 1, with the only difference that the Centrifugate of the low-nitrate rib extract solution is not added back to those ribs from which the Extract was obtained, but other ribs that same way were treated.

'" Example 4
Processing of tobacco leaves

1 kg of Maryland tobacco, in which 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 stored in 1250 ml at 50 ° C Occasionally immersed in water for 20 seconds. In doing so, the ones that are present in the superficial areas of the strips dissolved out water-soluble compounds. Since the nitrates and nitrites contained in the strips are mainly found in the If they are contained in superficial areas, most of them end up in the water. In the strip extract solution obtained in this way, which also contains other nitrogen compounds, the nitrogen compounds become microbiological degraded until the anions N.itrate_and Nitrite on one Content of 0.1 g per 1 are degraded.

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

Example 5 . - -

Obtaining the pure culture as in Example 1, Processing of tobacco leaves

250 g Burley tobacco leaves are mixed with 1250 ml at 50 ° C Water washed. The resulting tobacco extract solution is further treated like the rib extract solution according to the example 1 and the resulting low-nitrate tobacco extract solution is centrifuged off from the active sludge and added to the tobacco leaves again.

Example 6

T kg of tobacco waste, in which the nitrates and nitrites are to be reduced, is finely ground to a grain size of a maximum of 1 50> tim. 150 ml of active sludge, which are obtained according to Example 1 and are still in their exponential growth phase, are kept in this by adding the active sludge immediately together with 30 g of D-glucose, 9.6 g of NaCl, 675 g of KH ₂ PO ₄ and 35.25 g of Na ₃ PO ₄ ^ 2H ₃ O is added to 1500 ml of water. This gives a suspension of active microorganisms into which the tobacco powder is then stirred. The resulting pulp is aerated for 24 hours at 30 C under sterile conditions with an air pressure set of 3000 ml. min ""'in an Erlenmeyer container, which is closed with a porous stopper. The microorganisms in the tobacco powder build nitrates and nitrites and other nitrogenous compounds down to 1/10 of the original Immediately thereafter, 150 g of carboxymethyl cellulose are introduced into the paste

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and the pulp is spread into a 3 mm thick layer and dried to 15% moisture content. This stops the action of the microorganisms and solidifies the pulp into sheets of regenerated tobacco, which can be further processed. The regenerated tobacco obtained is low in nitrates and nitrites.

Example 7 · '

As in Example 6 with the only difference that, in accordance with Example 2, degradation takes place down to a nicotine content of 50 % .

Example 8

As in Example 1 with the only difference that the inoculated rib extract solution is held at 30 ° C. on a shaker 16 hours is incubated only 8 hours, so that the anions Nitrite and nitrate are degraded to a lesser extent than according to Example 1.

The low-nitrate rib extract solution obtained is then further treated as in Example 1 and added again to the ribs that were predried and washed out as in Example 1, 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 stems as in the ι example with the The only difference is that the inoculation is carried out with a pure culture of the bacterium Enterobacter aerogenes ATTCC 13048.

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Example 10

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

Example 11

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

Example 12

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

Example 13

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

Example 14

As in Example 8 with the only difference that the inoculation with a pure culture of the bacterium Enterobacter airborne ATTCC 13048 occurs.

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Claims (7)

FRG - P 28 PATENT CLAIMS "^ (η.) Process for refining tobacco by breaking down what is contained in tobacco Nitrates, nitrites and other nitrogen-containing compounds, characterized in that a culture of Microorganisms that need nitrogen and can meet their nitrogen needs through nitrate and nitrite breakdown under aerobic conditions Living conditions in their exponential growth phase ^ are mixed with a nutrient solution for this culture that is not essential for the microorganisms of this culture Has nitrogen source, under favorable aerobic living conditions for the microorganisms of this culture on the nitrates, nitrites and these accompanying other tobacco components to act until the Nitrates and other nitrogen-containing compounds are broken down to the desired residual content and that then immediately, After 24 hours at the latest, the effect caused by the microorganisms is broken off. 2. The method according to claim 1, characterized in that the culture of the microorganisms is brought to the action so intensively that the nitrates and nitrites are broken down within 6 to 16 hours to a minimum residual content and that the induced action of the microorganisms is terminated immediately as soon as the desired residual content is reached. 3. The method according to claim 1 or 2, characterized in that a pure culture of bacteria is applied, which is obtained by an aqueous smear of nitrate-containing leaves or rotten leaves are inoculated with a nutrient solution that has the nitrogen content required for incubation contains mainly in the form of nitrates, to pH 6.6 to pH 7.5 is buffered and at 25 to 35 ° C (degrees Celsius) for 6 to 16 hours by shaking under more sterile conditions 909834/0613 __ .; Aeration is incubated and then used as an active inoculum Inoculation of a fresh such nutrient solution is used, with which the incubation is then repeated and so on until the pure culture is formed. 4. The method according to claim 1 or 2, characterized in that a pure culture of the bacterium Enterobacter aerogenes from strain ATCC 13 048 is used. 5. The method according to claim 2, 3 or 4, characterized in that nitrates and nitrites and other water-soluble components be dissolved out with water from the tobacco to be refined that isolates the tobacco extract solution thus obtained inoculated with the microorganism culture and mixed with the nutrient solution under sterile intensive ventilation 6 to Is kept under favorable aerobic conditions for these microorganisms for 16 hours and then immediately afterwards the action of the microorganisms is terminated and that the contained in the low-nitrate tobacco extract solution Components of the solution are added to tobacco washed out with water. 6. The method according to claim 2, 3 or 4, characterized in that the tobacco to be refined is ground and slurried with water and mixed with the microorganism culture and nutrient solution is kept under sterile intensive ventilation for 6 to 16 hours under favorable aerobic living conditions for the microorganisms and that the action of the microorganisms is then immediately terminated in that the suspension is shaped into or onto foils and dried to a moisture content of 10 to 30%. SO 3834/0613 7. The method according to claim 5, characterized in that the active chi anun tobacco obtained is added and inactivated ν ± τά * _q; ■ · - '- / 0613