EP0575927B1 - Process for liming skins and hides - Google Patents

Abstract

The invention relates to processes for liming skins and hides using proteolytic enzymes in an aqueous alkaline liquor. The liming liquor has a pH within the range from 10 to 14 and contains, at one and the same time, thiourea dioxide and alkaline proteases having elastase activity.

Description

Field of the Invention

The invention relates to a method for liming skins and furs, the aqueous liming liquor containing alkaline proteases simultaneously with thiourea dioxide.

State of the art

In the course of the so-called water workshop in the production of leather, an alkaline process step, the liming, is usually used, which creates the conditions for depilation and brings about the necessary skin disintegration (Kirk-Othmer Encyclopedia of Chemical Technology 1st Ed., Vol. 8, 291 - 296, Interscience; Ullmann's Encyclopedia of Techn. Chemistry, 3rd ed. Vol. 11 , p. 560, 4th edition vol. 16 , pp. 118 - 119; F. Stather, tanning chemistry and tanning technology, Akademie-Verlag, Berlin 1967). In practice, you work with so-called sharpened liming, mostly a combination of calcium hydroxide and sodium sulfide. The implementation of the procedure depends on whether the hair should be destroyed or preserved. Attempts have been made to avoid the dangers associated with handling inorganic sulfide in various ways. Aside from avoiding conditions under which Hydrogen sulfide can be released, the interest has recently turned to the enzymatically supported liming process. (See E. Pfleiderer uR Reiner in HJ Rehm & G. Reed Ed., Biotechnology Vol. 6b , pg. 730 - 743, VCH, Weinheim 1988). Proteolytic enzymes [EC3.4] are mainly used, along with lipases [EC3.1.1.3] and amylases [EC3.2.1].

Task and solution

In order to reduce the content of the potentially dangerous sulfide in the wastewater, the proportional use of thio compounds, amines and hydrotropics has been proposed. With thio compounds alone, depilation can be performed. However, this fact does not mean that the problems are solved, since these substances also pollute the wastewater and lead to unpleasant odors. The enzymatic depilation is still only of limited importance, mainly for small animal skins and wool production. On the other hand, enzymatic hair removal in large cattle hides has not prevailed, primarily because of partly incomplete depilatory effects and because of damage to the collagen scar membrane or due to excessive breakdown of skin substance. The proportionate use of alkaline proteases in the liming along with small amounts of sulfides is also not unobjectionable. Thus, the sulfide content can be significantly reduced through the use of enzymes and you get very good surface yields with little grain, but the leather tends to loose grain, loose flame structure and a rough, sometimes nubucked grain pattern.

Some time ago, the use of thiourea dioxide (THDO) or formamidine sulfinic acid was proposed as a sulfide substitute (AT-PS 381 952, EP 197 918). This compound has a very high reduction potential compared to cysteine, so that in doses of 0.1-1% by weight, perfect depilation can be achieved together with calcium oxide or calcium carbonate. The compound is largely odorless and the degree of preservation of the hair is significantly better than that of a pure sulphide ash. In addition, the compound has a low toxicity to wastewater, since it is readily biodegradable. These advantages are offset by a relatively high price and the finding that the leather produced in this way does not have the optimal softness of the products treated in the conventional liming. These reasons are probably responsible for the fact that thiourea dioxide has so far not been successful in sole use. More recent proposals therefore result in the use of THDO together with hydrotropic and swelling-reducing substances, for example amines, in order to achieve the desired skin disruption with an appropriate amount of alkali (EP 306 474). In part, THDO is used in the post-scrubber without additional additives because of its bleaching effect, usually in amounts of 0.3 to 0.4% by weight, based on the pelvis weight.
In view of the prior art described, there was a need for a process which combines the advantages of the traditional liming process with the positive effects of the use of thiourea dioxide. Particular attention had to be paid to the ecological compatibility of the process and the active principles used.

It has now been found that the liming process according to the invention largely meets these requirements.

The present invention thus relates to a process for the liming of hides and skins, the aqueous liming liquors having a pH in the range 10-14, preferably 12-14, containing thiourea dioxide and alkaline proteases AP with elastase activity.
The liming liquors preferably contain 0.3 to 2% by weight, in particular 0.5 to 1% by weight, of thiourea dioxide together with an effective amount of one or more alkaline proteases AP. The liming process in the sense of the present invention summarizes the hair loosening, the actual liming, the skin disruption and, if necessary, the post-liming.
It is important that the presence of inorganic sulfide or sulfide ions and agents which develop sulfide under the reaction conditions can be dispensed with in the process according to the invention. The combination of THDO and alkaline proteases according to the invention is therefore excellently suitable for sulfide-free liming and / or post-liming as well as for low-sulfide liming / post-liming.

The alkaline proteases AP with elastase activity

To characterize elastases [EC3.4.21.11], their ability to hydrolyze elastin fibers of the aorta is used (W. Appel in HU Bergmeyer Ed. Methods of Enzymatic Analysis, 3rd Edition, Vol. I., pp. 1081-1085 Verlag Chemie 1974; J. Mandel in SP Cholowick and NO Kaplan: Methods in Enzymology, Vol. V, p. 665, Academic Press 1962).

Elastase preparations, even in crystallized form, must be regarded as inconsistent from the start; Even the purest preparations still contain a part of proteolytic, non-elastolytic activity. In structure and specific activity, the elastases seem to resemble trypsin and chymotrypsin.
The quantitative determinations are based on the (both proteolytic and mucolytic) degradation of the elastin. The main method of determination is the degradation of elastin, which is loaded with dyes such as orcin (or Congo red, dimethylaminonaphthalene sulfonic acid) or fluorescein.
The alkaline proteases AP to be used according to the invention with elastase activity are characterized by an optimum activity in the alkaline pH range, generally in the range pH 12 ± 2.
Although other sources are not to be excluded, microorganisms, in particular of the bacterial type, especially the bacilli, are currently the preferred starting materials.
Other examples include Flavobacterium elastolyticum, Chlortridium histolyticum, Staph. epidermis. In the case of preparations of alkaline proteases from Bacillus types, portions of 30-60% by weight of alkaline protease, 0.002-2% by weight of elastase in addition to neutral proteinase and collagenase (see USSR-PS 802 909, Chem. Abstr 94 , 148 340x).
Alkaline elastases have recently also been produced as products of genetic manipulation, for example by cloning the alkaline elastase gene from alkalophilic Bacillus and expression in Bacillus subtilis (cf. JP-OS 90 76 586, Chem. Abstr. 115 , 249 561c; Y.Ch. Tsai et al. Biochim. Biophys. Acta 1986 , 883 (3), 439-47, Appl. Environ. Microbiol. 1988 , 54 (12) 3156-61; Chem. Abstr. 110 ,, 110 535a; R. Kaneko et al. Japan. J. Bacteriol. 171 (9) 5232-36 (1989)). In the latter work the isolation of an alkaline elastase YaB from the alkalophilic Bacillus sp. YaB is produced extracellularly and expression of the gene in B. subtilis has been reported. Alkaline. Proteases with about 59% elastase activity were also obtained from an Aspergillus (A. versicolor 837) (see Chem. Abstr. 100 , 6 6 560x). Other sources are Pseudomonas types, e.g. P.aeruginosa (see A. Lazdunski et al. Biochimie 1990 , 72 (2-3) 147-56).

The determination of the elastase activity is carried out for the purposes of the present invention according to the method given in the experimental part. The units of elastase activity determined in the manner specified there are hereinafter referred to as EUgly elastase units. The definition is:
One elastase unit (EUgly) corresponds to the extinction of a µMol glycine by trinitrobenzenesulfonic acid determination; Analysis conditions: substrate is elastin, in buffer pH 8 and at 37 degrees C whereby the increase in extinction per minute is evaluated.
The activities of the active enzymes in the enzyme preparations AP are preferably in a certain ratio in the process according to the invention. This ratio is mathematically defined as follows: The protease activity of the alkaline protease [in Löhlein-Volhard units (LVE)] divided by a thousand times one Factor F gives the number of elastase activity in the units EUgly chosen for the purposes of the present invention (cf. experimental part).
According to the invention, the factor F is between 0.6 and 20, preferably 1 to 5.

The alkaline proteases [E.C.3.4.21] present in the enzyme preparations AP that can be used according to the invention are characterized in the usual way. (See Kirk-Othmer 3rd. Ed. Vol. 9, pp. 199-202, J. Wiley 1980; Ullmann's Encyclopedia of Industrial Chemistry Vol. A9, pp. 409-414, VCH 1987; L. Keay in "Process Biochemistry 17-21 (1971) These proteases, which mostly belong to the serine type, usually develop their optimum activity in a pH range of about 8-13. Bacterial proteases, in particular from Bacillus strains, are particularly mentioned, advantageously those which contain the elastase Bring activities from their origin, however, alkaline proteases of different origins can also be combined with one another, the elastase activity having to be introduced by appropriate addition.

Such alkaline proteases are, above all, those obtained from Bacillus strains, especially B.subtilis, but also B.formus, B.licheniformis, B.alcalophilus, B.polymixa, B.mesentericus, and also from Streptomyces strains such as S.alcalophilus called.
The cheapest working temperature with alkaline bacterial proteases (but in the present case must be clearly undercut) is generally 40 - 60 degrees C, with fungal proteases rather at 20 - 40 degrees C.
Alkaline fungal proteases include those from Aspergillus strains such as A.oryzae, from Penicillinum strains such as P.cyanofulvum or Paecilumyces persicinus. The activity of the alkaline fungal proteases is predominantly in the pH range 8.0 - 11.0.
The proteolytic activity of the alkaline proteases is customarily determined by the Anson hemoglobin method (cf. ML Anson J. Gen. Physiol. 22 , 79 (1939) or by the Löhlein-Volhard method (modified according to TEGEWA, cf. Leder, 22 , 121 - 126 1971).
One Löhlein-Volhard unit corresponds to the amount of enzyme that causes an increase in hydrolysis product in 20 ml casein filtrate, corresponding to an equivalent of 5.75 × 10 ^-3 ml 0.1 N NaOH. The protease activity to be used is generally between 1,000 and 60,000 LVU per kg skin, preferably between 2,000 and 14,000 LVU per kg skin.

Depending on the activity, the process according to the invention usually comes with amounts of proteases between 0.05 to 0.8% by weight, preferably about 0.1 to 0.3% by weight, as a rule of thumb when using an alkaline bacterial protease (Bacillus alcalophilus) ) with 4,000 LVE based on the weight of the hides and skins used.
According to the invention, 0.3-2% by weight, preferably 0.5-1% by weight, of thiourea dioxide are used together with the proteolytic enzymes AP. The fleet length is usually 100 to 120 wt .-% based on the weight of the hides and skins used.
The pH range of the liquor is advantageously adjusted using hydrated lime, but sodium hydroxide solution and / or soda can also be used proportionally.
To further improve skin disruption, agents known per se, such as, for example, organic amines, for example diethanolamine and / or hydrotropic substances such as, for example, urea, can also be used.

Execution of the procedure

As usual, fresh or salted raw goods are assumed. In general, a dirt switch and a main switch are carried out for the pretreatment (US Pat. No. 4,344,762). The main switch, as is customary in the business, is carried out using suitable proteases and / or surfactants at a pH of 9-10 over a period of 4-6 hours.
The main switch fleet is typically drained and a new bath is continued. In general, the enzymatic reaction is carried out in the temperature range 20-28 degrees C., preferably at 26 degrees C. The liquor containing the enzymes and the thiourea dioxide, which is adjusted to an alkaline pH, especially in the range 10-13, is left in a conventional reaction vessel, for example a mixer, tanning drum, etc., with agitation, for example, for a sufficient period of time, as a rule of thumb there are approximately 90 Called minutes, act on the hides and skins until they are largely hair-free.
Then it can be post-alkalized with a little alkali, for example 0.2% by weight of a 50% sodium hydroxide solution, preferably with agitation for about 30 minutes. This is followed by a longer treatment phase, expediently with short-term movement / longer rest, for example in rotation: 1 minute movement, 59 minutes rest, which is carried out, for example, over 18 hours. The fleet is then drained. The hair turns out to be less destroyed than when using a conventional sulphide limescale. Washing is advantageous, for example twice with 200% water at 25 degrees C over 15 minutes.
Further processing can be carried out in a conventional manner, for example in the sequence pickling / decalcification / pimples / chrome tanning.

Beneficial effects

The process according to the invention permits the production of remarkably soft leather, it being particularly emphasized that despite the use of degrading enzymes there is generally a completely intact scar pattern. Overall, the result can be regarded as very surprising, since the nubucking to be expected when using the enzyme in the liming occurs less than the expected loose grain in the flames.

By using the enzyme in the context of the method according to the invention, the required amount can be used Significantly reduce thiourea dioxide. The combination of thiourea dioxide and alkaline protease in the liming thus enables an ecologically extremely favorable liming process, which combines high leather quality with sufficient application safety. The ecological advantage lies primarily in the good hair preservation and thus the lower COD pollution in the wastewater as well as in the avoidance of any use of sulfide.

The following examples serve to illustrate the invention.

EXAMPLES Examples 1-4: Raw goods:

1 t salted or fresh cowhide, weight class 30 - 39 kg (black and white).

Pretreatment:

Dirt diverter, main diverter is normally used by using tensides at pH 9-10 for 4-6 hours. The main switch fleets are drained and all examples continue to work in a new bath.

All percentages represent% by weight based on the salt or green weight of the skins.

Example 1: Raw goods:

1 t salted beef skins

150,0 %

Wasser, 26 Grad C

3,5 %

Kalk

0,8 %

Thioharnstoffdioxid

0,3 %

proteolytisches Enzym, z.B. aus Bacillus alcalophilus, pH-Wirkungsoptimum
pH 10 - 13, 4 000 LVE-Einheiten, Elastasewert 6,4 (F = 1,6)
90 Min. bewegen bis Häute weitgehend haarfrei

0,2 %

Natronlauge 50 %ig
30 Min. bewegen
anschließend weitere 18 Stunden behandeln (1 Min. bewegen, 59 Min. ruhen).
Flotte ablassen (Haare sind geringer zerstört als bei konventionellem Sulfid/Kalk-Äscher)
2 x waschen mit je 200 % Wasser, 25 Grad C, 15 Min.

betriebsübliche Weiterarbeit mit Beize/Entkälkung/Pickel/ Chromgerbung.Liming:

150.0%

Water, 26 degrees C.

3.5%

lime

0.8%

Thiourea dioxide

0.3%

proteolytic enzyme, e.g. from Bacillus alcalophilus, optimum pH effect
pH 10 - 13, 4,000 LVE units, elastase value 6.4 (F = 1.6)
Move for 90 minutes until the skin is largely hair-free

0.2%

Sodium hydroxide solution 50%
Move for 30 minutes
then treat for another 18 hours (move 1 min., rest 59 min.).
Drain the liquor (hair is less destroyed than with conventional sulphide / limestone liming)
Wash twice with 200% water, 25 degrees C, 15 min.

customary further work with pickling / decalcification / pimples / chrome tanning.

Example 2 Raw goods:

1 t fresh cowhide

100,0 %

Wasser, 26 Grad C

1,0 %

Kalk, 90 Min. bewegen, pH 12 - 12,5

1,0 %

Thioharnstoffdioxid

0,3 %

proteolytisches alkalistabiles Enzym (z.B. aus Bacillus alcalophilus)
pH-Wirkungsoptimum pH 10 - 13, 4 000 LVE,
Elastasewert 6,0 (F = 1,5)
90 Min. bewegen, pH 12 - 13, Häute sind haarfrei und gut in ihrer Struktur erhalten;
Haar kann durch Umpumpen der Flotte über ein Sieb abgetrennt werden.

+ 50,0 %

Wasser, 26 Grad C

2,5 %

Kalkhydrat
10 Min. bewegen

0,2 %

Natronlauge 50 %ig
weitere 15 Stunden behandeln (Automatik: 2 Min. bewegen, 58 Min. ruhen): Flotte ablassen
2 x waschen mit je 250 % Wasser, 26 Grad C, 15 Min.

Weiterarbeit betriebsüblich.Liming (hair preserving):

100.0%

Water, 26 degrees C.

1.0%

Lime, move for 90 minutes, pH 12 - 12.5

1.0%

Thiourea dioxide

0.3%

proteolytic alkali-stable enzyme (e.g. from Bacillus alcalophilus)
Optimal pH effect pH 10 - 13, 4,000 LVE,
Elastase value 6.0 (F = 1.5)
Move for 90 minutes, pH 12 - 13, skins are hair-free and their structure is well preserved;
Hair can be separated by pumping the liquor over a sieve.

+ 50.0%

Water, 26 degrees C.

2.5%

Hydrated lime
Move for 10 minutes

0.2%

Sodium hydroxide solution 50%
Treat for another 15 hours (automatic: move for 2 minutes, rest for 58 minutes): drain the liquor
Wash twice with 250% water, 26 degrees C, 15 min.

Continued work customary.

Example 3 Raw goods:

1 t salted beef skins

150,0 %

Wasser, 26 Grad C

3,5 %

Kalkhydrat

0,4 %

Natriumsulfhydrat, 72 %ig
30 Min. bewegen

0,4 %

Thioharnstoffdioxid

0,1 %

proteolytisches, alkalistabiles Enzym (z.B. aus Bacillus alcalophilus), 4 000 LVE, Elastasewert 6,8 (F = 1,7)
60 Min. bewegen bis Häute haarfrei

+ 0,3 %

Natronlauge, 50 %ig weitere 18 Stunden bewegen (Automatik: 2 Min bewegen, 58 Min. ruhen).
Flotte ablassen
2 x waschen mit je 150 % Wasser, 25 Grad C, 15 Min.

Weiterarbeit betriebsüblich.Low sulphide liming:

150.0%

Water, 26 degrees C.

3.5%

Hydrated lime

0.4%

Sodium sulfhydrate, 72%
Move for 30 minutes

0.4%

Thiourea dioxide

0.1%

proteolytic, alkali-stable enzyme (e.g. from Bacillus alcalophilus), 4,000 LVE, elastase value 6.8 (F = 1.7)
Move 60 min. Until skins are hair free

+ 0.3%

Sodium hydroxide solution, move 50% for another 18 hours (automatic: move 2 minutes, rest 58 minutes).
Drain the fleet
Wash twice with 150% water, 25 degrees C, 15 min.

Continued work customary.

Example 4

Conventional sulphide / lime liming and post-liming with active ingredient combination according to the invention for the production e.g. especially soft furniture leather with high color levelness.

Raw goods:

1 t salted beef skins

150,0 %

Wasser, 26 Grad C

2,0 %

Kalkhydrat

0,9 %

Natriumsulfhydrat
20 - 30 Min. bewegen

1,0 %

Kalkhydrat, 20 Min. bewegen

0,4 %

Natriumsulfid, 60 %ig
30 Min. bewegen
dann Automatik: 2 Min. bewegen, 58 Min. ruhen insgesamt 15 Stunden
Flotte ablassen, 2 x waschen, Häute entfleischen und spalten auf 1,8 - 2 mm.

Liming:

150.0%

Water, 26 degrees C.

2.0%

Hydrated lime

0.9%

Sodium sulfhydrate
Move for 20-30 minutes

1.0%

Hydrated lime, move for 20 min

0.4%

Sodium sulfide, 60%
Move for 30 minutes
then automatic: move for 2 minutes, 58 minutes rest for a total of 15 hours
Drain the liquor, wash 2 times, remove the skins and split to 1.8 - 2 mm.

150,0 %

Wasser, 26 Grad C

1,0 %

Kalkhydrat

0,3 %

Thioharnstoffdioxid

0,1 %

proteolytisches, alkalistabiles Enzym (z.B. aus Bac.alcalophilus), 4 000 LVE, Elastasewert 9,2 (F = 2,3)
20 Min. bewegen, dann weitere 6 Stunden: 2 Min. bewegen, 58 Min. ruhen.

Flotte ablassen, waschen und betriebsüblich
weiterarbeiten.Post-purifier:

150.0%

Water, 26 degrees C.

1.0%

Hydrated lime

0.3%

Thiourea dioxide

0.1%

proteolytic, alkali-stable enzyme (e.g. from Bac.alcalophilus), 4,000 LVE, elastase value 9.2 (F = 2.3)
Move 20 minutes, then another 6 hours: move 2 minutes, rest 58 minutes.

Drain the fleet, wash it and normal use
continue working.

Determination of the elastase activity of the enzymes used according to the invention.

Principle:

An elastin suspension of pH = 8 is incubated with enzyme at 37 ° C. for 2 hours, terminated by filtering off the substrate, stained with TNBS and measured at 420 nm.

Definition:

1 unit of elastase corresponds to the amount of enzyme that causes staining with TNBS equivalent to 1 µmol of glycine per minute in an elastin suspension under the specified standard conditions.

Reagents:

Elastin (Sigma Lot 71 F-8020; No. E-1625)
Boric acid pa (= per analysis)
Trinitrobenzenesulfonic Acid (TNBS)
Glycine

Equipment:

Shake thermostat: 37 degrees C.
Water bath: 50 degrees C.

Solutions: 1. 0.1 M borate buffer, pH = 8.0

A solution of 6.2 g of boric acid pa is adjusted to pH = 8.0 with 1N NaOH and with dist. Make up to 1 liter of water.

2. TNBS reagent

Approx. 800 ml dist. 6.2 g of boric acid pa are added to water with stirring and the pH is adjusted to 8.0 with 1N NaOH. Add 240 mg TNBS, adjust the pH if necessary and fill with distilled water. Water to 1 l.
The TNBS reagent is best kept in a brown bottle and has to be prepared daily.

Reaction: Main value:

250 mg of elastin are weighed into a 50 ml narrow-neck Erlenmeyer flask with a ground glass stopper and mixed with 10 ml of 0.1 m borate buffer. The flask is preheated in the shaking thermostat for 10 min. After adding 1 ml of enzyme solution, the mixture is mixed well and the flask is returned to the shaking thermostat. Temperature: 37 degrees C.

The reaction is stopped by filtering the reaction mixture through a pleated filter after exactly 2 hours. The fragments are immediately stained using the TNBS method .

TNBS: Response:

100 μl of the sample are added to 8 ml of TNBS reagent and the test tube is left for 25 min. stand in a water bath at 50 degrees C. After exactly 25 min. the test tube is 5 min. placed in ice water and immediately afterwards the absorbance measured at 420 nm.

Blank value:

The enzyme solution is only added after the second hour of the reaction time. The further treatment is the same as for the main value, i.e. begins with the stopping.

Creation of a calibration curve with glycine:

The staining of glycine with TNBS is measured and in addition µmol of glycine against O.D. applied.

Method:

3.75 g of glycine are dissolved in 100 ml of distilled water, 250 ml of which are removed and diluted to 500 ml. 100 µl are removed and stained using the TNBS method. The measurement is carried out at 420 nm. Additional weights are selected accordingly. The following table gives an example of a calibration curve: Weigh-in (g) µmol / ml OD at 420 nm 3.75 / 100 - 0.25 / 500 0.25 0.30 3.75 / 100 - 0.25 / 250 0.5 0.058 3.75 / 100 - 0.50 / 250 1.0 0.110 3.75 / 100 - 1.00 / 250 2.0 0.232 3.75 / 100 - 1.00 / 200 2.5 0.313 3.75 / 100 - 1.50 / 200 3.75 0.496

The blank value (O.D. of TNBS reagent + 10 l distilled water) was previously subtracted from all glycine values.

Activity calculation:

$$\text{Elastase Einheit/mg =}\frac{\text{µmol Glycin (auf Eichkurve abgelesen) x 91,7}}{\text{C in mg}}$$

The extinction difference of the activity determination (main value minus blank value) is converted from the calibration curve into µmol glycine, which results in the elastase units $$\text{Elastase unit / mg =}\frac{\text{µmol of glycine read on calibration curve x 11 x 100}}{\text{120 min x conc. enzyme}\left(\text{C.}\right)\text{in mg}}$$

$$\text{Elastase unit / mg =}\frac{\text{µmol of glycine (read on calibration curve) x 91.7}}{\text{C in mg}}$$

Example:

Weighing enzyme (g) 0.5 / 50 - 5/50 from this concentration / ml 1 mg / ml Main value (420 nm) 0.408 Blank value 0.053 Extinction 0.355

From the glycine calibration curve: An OD of 0.355 corresponds to 2.79 mol of glycine. $$\text{Elastase units / mg =}\frac{\text{2.79 x 91.7}}{\text{1}}\text{=}\underline{\underline{\text{255.8}}}$$

Claims (6)

1. A process for liming skins and hides using proteolytic enzymes in an aqueous alkaline float, characterised in that the liming float with a pH-value in the range of 10-14 comprises both thiourea dioxide and alkaline proteases AP with elastase activity.
2. A process according to claim 1, characterised in that the liming float comprises 0.3 to 2 wt.% of thiourea dioxide.
3. A process according to claims 1 and 2, characterised in that the liming float, in addition to alkaline protease [E.C.3.4.21], comprises an active amount of alkaline elastase [E.C.3.4.21.11].
4. A process according to claims 1 to 3, characterised in that the liming float has no added sulphide.
5. A process according to claims 1 to 4, characterised in that the alkaline protease is a bacterial protease.
6. A process according to claim 5, characterised in that the alkaline bacterial protease was obtained from Bacillus alcalophilus.