WO2012052405A2

WO2012052405A2 - Method for the preparation of cationic surfactants

Info

Publication number: WO2012052405A2
Application number: PCT/EP2011/068116
Authority: WO
Inventors: Xavier Rocabayera Bonvila; Joan Seguer Bonaventura; Francisco Javier Rodriguez Martinez; Eva Piera Eroles
Original assignee: Laboratorios Miret, S.A.
Priority date: 2010-10-21
Filing date: 2011-10-17
Publication date: 2012-04-26
Also published as: WO2012052405A3

Abstract

The invention relates to a method of the preparation of a cationic surfactant, derived from the condensation of fatty acids and esterified dibasic amino acids, having the formula (I): having a mean particle diameter of less than 300 μm; by preparing the cationic surfactant with a remaining content of water less than 5% by weight in a first step; and milling this dried material in the second step.

Description

METHOD FOR THE PREPARATION OF CATIONIC SURFACTANTS

This invention relates to a method for the preparation of ethyl ester of the lauramide of the arginine monohydrochloride.

In the art it is known to use cationic surfactants as antimicrobial agents in the food, cosmetic and pharmaceutical industry and also in medical devices. All these cationic surfactants contain at least one alkyl chain, which forms the basis of the hydrophobic section of the molecule.

A particular group of cationic surfactants are the arginine derivatives, such as the lower alkyl esters of mono-N-higher aliphatic acyl arginine. This group is known in the art to have good antiseptic, medicinal, preservative, bactericidal, germicidal and fungicidal properties without causing significant irritation and skin troubles. The common chemical structure of the arginine derivatives is as follows (1 ):

The antimicrobial activity of this specific type of cationic surfactants of formula (1 ) is highly unpredictable because the fatty acids (RCO) and the alkyl radical (Ri) can vary in structure and chain length. Furthermore, the R groups may be saturated or unsaturated, cyclic or acyclic and branched or non-branched. Thus, the distribution of chain lengths constitutes a "fingerprint" on the final compound. In this sense, there is a considerable number of possible compounds included in the arginine derivatives like lower alkyl esters of mono-N-higher aliphatic acyl arginine. When they were used in food products or cosmetic products, unexpected effects could be observed.

Extensive studies have been performed on the structural, physicochemical and biological properties of long chain N"-acyl amino acids. In general these surfactants show a rich multifunctionality with a relationship between their structure and properties. Several studies have been conducted to assess the influence of several structural modifications, for example the straight chain length of fatty acid, or the nature of the R substituent.

For the specific case of the cationic surfactant derived from lauric acid and arginine, the ethyl ester of the lauramide of the arginine monohydrochloride, hereafter referred to as LAE has been found to be a highly effective substance against microorganisms (i.e.: bacteria, virus, yeasts and moulds). Nowadays, LAE is a well-known antimicrobial agent with low toxicity to human beings and good efficacy when it is applied as a preservative alone or in combination with other substances in food and cosmetic matrices or in medical devices.

LAE has the chemical structure of formula (2) displayed hereafter.

Several patents of Laboratorios Miret, S.A. (LAMIRSA) describe the manufacturing process to obtain LAE, i.e.: EP 0 749 960 B1 and EP 1 294 678 B1 . These patents cover different process to obtain a cationic surfactant of the formula (1 ) and specifically LAE (formula 2). According to these patents the crude reaction product obtained at the end of the synthesis is filtered through a filter press in order to isolate the final product obtained via the reactions described in these patents. Once the filtration process is finished, the resulting product is a white solid with a yield of 80-85% w/w with respect to the product initially expected.

The LAE that has being obtained at the LAMIRSA premises contains an important content of water. If the final product is only filtered, the range of water content is 1 1 .5-20%, and if the filtered product is dried by usual methods, such as under control heat, the final product LAE has a range of water content between 5 - 6 %. The particles of LAE achieved at the end of the drying process display a mean particle diameter of approximately 2000 μΐη.

In general, the standard methods for the preparation of the cationic surfactants of the formula (1 ) lead to a final product with particles having a mean particle diameter of around 2000 μΐη.

According to the present invention the cationic surfactants according to formula (2) shall have a mean particle diameter of less than 300 μΐτι. In the constant research to improve the efficacy of LAE, it has surprisingly been found that the extraction of the water to reduce its content below 5% is extremely important. This reduction of water content allows decreasing the mean particle diameter of the cationic surfactant.

It requires considerable efforts to obtain LAE of formula (2), with a remaining water content of approaching zero. Typically, in drying, one does not physically or chemically change the composition of the product, other than by removing the moisture. The main difficulty to reduce the water content of LAE is its low melting point which is around 50°C and also its surfactant properties such as the low surface tension. In this sense, while trying to eliminate the content of water with conventional methods like heating LAE above 30°C, there is observed a change of the LAE properties providing a pasty type of LAE, similar to a gum. This texture makes it considerably difficult to apply LAE in the matrices with the intention to protect them against microbial growth.

With the current manufacturing methods for LAE, the presence of one molecule of water per molecule of the surfactant leads to an amount of water relative to the total weight of the surfactant of 4.3 % by weight, this content of water is known as intramolecular water of LAE. At the end of the drying process the usual content of water is between 5 to 6%, thus there is a superficial content of water from 0.7-1.7% and 4.3% of intramolecular water.

The particle diameter of LAE with the current manufacturing process is ca. 2000 μηη and it is not possible to reduce its particle diameter to less than 300 μηι with a milling process after drying LAE with conventional methods because the content of water in LAE is still between 5 - 6% and this forms a sticky paste.

The conventional drying methods can be static or dynamic. Drying LAE with a dynamic process consists of shaking the product in a vacuum process below 30°C. The result obtained was a sticky paste produced by the continuous agitation of LAE. Thus, the subsequent process to grind LAE is impossible to be carried out.

With a static process the LAE is dried through an electric oven with heated air below 30 °C. After a long period of drying the final product which was obtained was a LAE with 5 - 6% content of water, which product forms a sticky paste during the grinding process.

Surprisingly, it was observed that through the reduction of the superficial water content leading to a LAE with less then 5% of water, the formation of the sticky paste of LAE was avoided and it was possible to grind LAE up to 300 μιη particle diameter with a milling process. The combination of the use of the TRI-CHOP equipment with a milling process was the best manner allowed to obtain a LAE with less than 5% of water content and a diameter of particle inferior to 300 μΐη.

The preferred remaining water content during the drying procedure was less than 5% by weight, preferably less than 4.5% by weight, more preferably less than 2.5% by weight and most preferably no water at all (0%). The method of the preparation of the cationic surfactants with a mean particle diameter of less than 300 μνη provides for an additional step of reducing the particle diameter of the product which is obtained in the conventional manner of preparation.

The mean particle diameter of the cationic surfactants described in the present application is determined on the basis of the methods given in UNE 55-550-78 and ASTM E 1 1 -95 (corresponding to ISO 3310-1 ).

To reduce the particle diameter of LAE (less than 2000 μηη and with 5-6% of water content), the conventional methods of size reduction and sieving have been tried out. However, it turned out that simple milling and sieving was not possible with this particular class of compounds. The manipulation of the compound led to the generation of a pasty type of product with loss of the detection of individual particles.

This reduction of the particle diameter can be conducted by subjecting the particles which have been achieved to any manner described in the art. It is a preferred method of using milling and more preferably a colloid mill for reducing the diameter of the particles. However, it has been surprisingly observed, that the size reduction of the particles, preferably the size reduction with the colloid mill method, is more favourable when the size reduction is conducted using particles of LAE which contain a particularly low concentration of water. Besides, this reduction of water content improves the stability of formulations of LAE because the absence of water lowers the hydrolysis of LAE.

It is for instance highly suitable to remove water from LAE with a particular special equipment developed to reach very low water concentrations. Such equipment is sold under the brand name TRI-CHOP by the company Lleal S.A. (Granollers, Spain) in different sizes. This particular equipment has been developed applying the most advanced design technology with the aim to optimize drying processes.

The drying process using the TRI-CHOP equipment is intended to remove moisture from the cationic surfactant to obtain a "dry" final product. This does not imply that the product will have no remaining water at all. Dry in the sense of the present invention means that the product contains less than 5% by weight, preferably less than 4.5% by weight, more preferably less than 2.5% by weight and most preferably no water at all (0%).

The best conditions for drying are a pressure range of lower than 50 mbar preferably lower than 20 mbar and most preferably lower than 5 mbar, and a temperature range of 10- 35°C, preferably 25-30°C. The duration of drying depends on the amount of the product and the final result to be achieved in terms of the amount of water.

Once the cationic surfactant is dried, meaning in the sense of the present invention once a final water content inferior to 5% by weight has been attained, the product is passed through a mill, preferably a colloid mill, in order to reduce the mean particle diameter to a diameter lower than 300 pm. It is in general very difficult to reach a more "dried LAE" with a content of water less than 5% and this purpose has been surprisingly achieved with the TRI-CHOP equipment. This new "dried LAE^" is the only type which is suitable to have its particle diameter reduced to less than 300 μιη through a milling process. The amount of water in the compounds of the formula (1 ) is usually determined by the Karl Fischer method.

The use of LAE with small particles is new. Because of its high reactivity due to the large surface to volume ratio, LAE with small particle diameter plays a crucial role in inhibiting bacterial growth in solid media.

The antimicrobial activity of LAE is influenced by the dimension of the particles, the smaller the particles, the greater the antimicrobial effect.

The LAE with the reduced mean particle diameter according to the present invention can be used in any application which has been described for the cationic surfactants before. In particular the cationic surfactants with the reduced mean particle diameter can be used in solid food applications, such as in the preservation of meat products, like for instance meat, poultry products, fish, crustaceans, vegetables, greens, emulsions, sauces, confectionery, gums, bakery, pre-cooked meals, ready-to-serve meals, dairy products, egg-based products, jams, jellies, beverages, juices, wines and beers.

LAE according to formula (2) with the reduced mean particle diameter prepared according to the present invention can be used in any application related with cosmetic products (for instance, creams, toothpaste and deodorants).

In all of these applications the small sized LAE turned out to display a much greater activity than the regularly sized corresponding products.

The invention is illustrated through the examples provided hereafter. EXAMPLES.

Example 1 .

PREPARATION OF LAE WITH SMALL PARTICLE DIAMETER

30 kg of LAE obtained following the method of EP 1 294 678 B1 with a content of water of 19.5%, determined by Karl Fischer method. The mean particle diameter was 2,000 pm determined by UNE 55-550-78 and ASTM E 1 1 -95. For drying it was used the TRI-CHOP equipment with the following conditions: a pressure of 10-20 mbar, and a temperature range of 25-30°C. The initial turning speed was 15 rpm during 4 hours, and then it was increased up to 30 rpm for further 20 hours.

This LAE had a water content of 4.5% determined by Karl Fischer method and 89.8% of active ingredient LAE (JECFA 71 ^th Monograph). There is no change of the mean particle diameter during this drying process.

For the milling process it was used a colloid mill from MICRON PROCESS with the following conditions: 2,000 rpm in the separator and 4,000 rpm in the mill. The flow rate was 24 kg/h.

The final quantity of LAE was 25.5 kg. The mean particle diameter was 270 μιη determined by UNE 55-550-78 and ASTM E 1 1-95.

Example 2.

IMPROVED STABILITY

This example compares the stability of three different samples of LAE in a liquid solution along the time and at two different temperature conditions (21 °C and 40°C). Each sample of LAE was obtained with different methods and consequently the content of water in each LAE was also different:

Sample A: LAE obtained at the end of the filtered process following EP 1 294 678 B1 . This LAE had a water content of 20% and 72.3% of the active ingredient LAE. The mean particle diameter was 2000 μΐη.

Sample B: LAE obtained after filtering following EP 1 294 678 B1 and then drying under heat (usual method). This LAE had a water content of 5.8% and 88.8% of active ingredient LAE. The mean particle diameter was 2000 μΐη.

Sample C: LAE used was the one obtained in Example 1 . This LAE had a water content of 4.5% and 89.8% of active ingredient LAE. The mean particle diameter was 270 μΐη.

To compare the stability of LAE in a liquid solution, samples A, B and C were dissolved in a food grade solvent. In each case the final liquid formulation had to contain 20% of active ingredient of LAE and ca. 80% of glycerin as the solvent. The stability was determined according to the following equation with which the percentage of degradation of LAE was calculated along the time:

% degradation = I^gj^ ^{~ [LAE}^< L_{x m}

On the basis of the calculation with the above formula the figure 1 illustrates the percentage of degradation of LAE in time (expressed in weeks) for LAE with 20%, 5.8% and 4.5% of water content at 21 °C. The results surprisingly show, that reducing the content of water in LAE results in a better stability of the molecule.

The figure 2 illustrates the behavior of LAE with the different content of water when it is submitted at higher temperature, i.e. 40°C.

The data in figure 2 surprisingly show how under extreme conditions, where LAE is stored at 40°C, the presence of water affects the stability of LAE increasing its degradation and how decreasing the content of water to below 5% guarantees a better stability along the time.

Example 3.

APPLICATION IN SHREDDED CHEESE TREATMENTS SHREDDED CHEESE

Treatments: Control, 200 mg/kg and 400 mg/kg of LAE in the final product.

LAE with small particles (sp LAE): a mean particle diameter of 270 μηι and 4.5% water content obtained with the TRI-CHOP equipment and a milling process, as described in Example 1 .

Standard LAE (Std LAE) with a mean particle diameter of 2000 μηι and 5.8% of water content obtained from the press filter following EP 1 294 678 B1 and dried under control heat. Procedure:

a. - Shred the cheese; weigh the cheese (96 g).

b. - Blend the shredded cheese with 4 g of a blend of LAE (Std or sp) with Starch.

Dose levels for each LAE (Std and sp): 200 mg/kg and 400 mg/kg.

Starch was C^*Gel 03401 (Corn starch) by Cargill.

Once applied to the product, the treated sample was made into individuals samples of 20 g each.

HPLC analysis was conducted at 1 h after application, 24 h, 48 h, 5 days and 15 days with one of the samples.

With the other samples, inoculation of L. monocytogenes (10⁵ cfu/g) was carried out. The control samples are the shredded cheese with just starch. Microbiological analysis at times: immediately after application (1 hour), 24 h, 48 h, 5 days and 15 days. Temperature control: 4 °C (39°F).

Each sample was stored in a plastic bag in vacuum.

RESULTS

a) Microbiological results

Table 1

Table 2

Example 4.

APPLICATION IN CHEWING GUM Chewing gum is a particular food matrix with increasing interest to apply LAE to achieve better breath freshening chewing gums that prevents the halitosis.

RESULTS: Antimicrobial activity of LAE on the biofilm with five bacteria and a yeast:

The results obtained with a control biofilm were compared with a biofilm treated with 0.5% standard LAE (mean particle diameter ca. 2000 μΐη) obtained with the press filter and then dried under control heat and 0.5% small particles LAE (270 μΐη) obtained with the TRI-CHOP equipment and a milling process.

The results of the following table were expressed as log of CFU : Table 3: Antimicrobial activity of LAE versus control

0.5% small

0.5% LAE

Microorganism Control particles LAE

(Std LAE)

(sp LAE)

A. naeslundii 8.3 2.5 <2.0

V. dispar 8.1 5.0 4.1

F. nucleatum 5.1 2.1 <2.0

S. sobrinus 8.6 6.1 3.8

S. oralis 9.0 6.1 4.3

C. albicans 5.1 2.1 <2.0

Limit of detection: 2.0 The results demonstrate the efficacy of the antimicrobial effect of standard LAE in a biofilm and a further improvement when LAE small particles are used. The surprising decrease of the number of colonies of those microorganisms responsible to provoke halitosis is thanks to the presence of LAE in the chewing gum. Nevertheless, the most significant reduction on the number of colonies is observed in the chewing gum treated with the small particles of LAE.

Claims

1 . Method of the preparation of a cationic surfactant, derived from the condensation of fatty acids and esterified dibasic amino acids, having the formula:

having a mean particle diameter of less than 300 μΐη;

by preparing the cationic surfactant with a remaining content of water less than 5% by weight in a first step; and

milling this dried material in the second step.

The method of claim 1 , whereby the remaining content of water is 4.5% by weight or less.

3. The method of claim 1 , whereby the remaining content of water is 2.5% by weight or less.

4. The method of any of the claims 1 to 3 to be applied in food products, cosmetic products and medical devices.

5. The method of any of the claims 1 to 4 whereby the preferred food products are solid matrices.