JP2006070231A - Biodegradable liquid detergent composition - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a detergent composition containing an acid type sophorolipid chemically stable even in a liquid. <P>SOLUTION: The biodegradable liquid detergent composition comprises sophorolipid, wherein the sophorolipid contains at least 90% sophorolipid (acid type). The sophorolipid (acid type) is obtained by mixing sophorolipid with an alkali at room temperature. The sophorolipid is practically 100% acid type sophorolipid. The composition contains 0.01-20 wt.% acid type sophorolipid. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a biodegradable liquid detergent composition. More specifically, the present invention relates to a cleaning composition suitable for a cleaning process that includes acid sophorolipid as a surfactant and requires low foaming properties.

  Currently, the types of surfactants are very diverse, and various surfactants are selected depending on how the final product is used. However, from the consumer's point of view, there are only two alternatives: synthetic surfactant-containing products or soap-containing products. Consumers who select products from the viewpoint of biodegradability and safety choose soaps over synthetic surfactants, but soaps are less detersive than synthetic surfactants, so use them all at once It is pointed out that although the amount is large, the biodegradability is high, but the load on the environment is large. In other words, a third surfactant (next-generation surfactant) that has the same biodegradability and safety as soap and the same performance (detergency, hard water stability, etc.) as a synthetic surfactant has been put to practical use. It is desired.

  In the field of detergents, jet cleaning, which removes dirt from objects to be cleaned by using water pressure for cleaning, has attracted attention as a new cleaning method. It is applied to infectors. If a normal surfactant with high foaming properties is used as a cleaning agent for this jet cleaning, the jet water pressure decreases due to the large amount of foam generated, and a satisfactory cleaning effect cannot be obtained. Overflow from the machine and washing tank causes trouble in the washing process. Therefore, it is necessary to use a surfactant having a low foaming power, that is, a low foaming property, for jet cleaning.

  In order to perform jet cleaning, a method of adding an antifoaming agent (typically a silicon-based antifoaming agent) was studied, but satisfactory results were not obtained in terms of cleaning power and defoaming power.

  At present, cleaning agents containing block polymer type nonionic surfactants are mainly used for jet cleaning. This block polymer type nonionic surfactant contains ethylene oxide (EO), propylene oxide (PO) and the like in the molecule, and has a low foaming power, that is, low foaming property. However, the greatest problem is that the biodegradability in the environment is extremely poor (Journal of The American Oil Chemist's Society, 65, 1669-1676 (1988)).

  In order to improve biodegradability in the environment, block copolymers in which the degree of polymerization of PO is changed, block polymers having terminal alkyl modification, and the like have been synthesized, but the problem has not been solved.

  Biosurfactants, which are biologically derived surfactants, are attracting attention as next-generation surfactants because of their high biodegradability and safety. However, since it is more expensive than synthetic surfactants, only application research to cosmetics, pharmaceuticals, and the like with high added value can be seen. In addition, there is very little research and development in the field of detergents where the products themselves are inexpensive.

  An example of using a biosurfactant as a cleaning agent is sophorolipid (lactone type) in which sophorolipid is blended as a low-foam surfactant and the composition of the sophorolipid is 35% or more, preferably sophorolipid (lactone type) and sophorolipid. (Acid type) in a ratio of 35:65 to 90:10, and as washing auxiliary components, enzymes, oxygen bleaches, bleach activators, alkali agents, water softeners, fluidity modifiers and neutrals A powder low-foaming detergent composition comprising anhydrous sodium sulfate is already known (Japanese Patent Laid-Open No. 2003-13093, Patent Document 1).

  However, sophorolipid (lactone type) containing sophorolipid is a very excellent surfactant that satisfies biodegradability, high detergency, and low foaming properties. Is chemically unstable, so its use is very limited, and it has been very difficult to develop a product as a third surfactant after soap and synthetic surfactant. For example, when blended as a liquid alkaline detergent, the lactone moiety is hydrolyzed.

In other words, if a chemically stable sophorolipid is manufactured at a low cost and has performance (detergency, foamability, compatibility with each component, storage stability, etc.) that can withstand actual use as a cleaning agent, The use of detergents formulated with it will become much broader than it currently is, and development of products that incorporate biosurfactants will proceed.
JP 2003-13093 A

  An object of the present invention is to provide an acid-type sophorolipid-containing cleaning composition that is chemically stable even in a liquid.

  As a result of intensive studies on the performance of the sophorolipid obtained by allowing the sophorolipid obtained by fermentation to stand under alkaline conditions, the present inventors have completed the present invention. The inventors of the present invention are that the sophorolipid obtained by fermentation becomes an acid-type sophorolipid only by standing under alkaline conditions, the obtained acid-type sophorolipid is chemically stable, exhibits low foaming property, and The present inventors have found that it has a high detergency and have completed the present invention.

  That is, the biodegradable liquid detergent composition of the present invention is a biodegradable liquid detergent composition containing sophorolipid, and the sophorolipid contains at least 90% or more of sophorolipid (acid type). This achieves the above object.

  In one embodiment, the sophorolipid (acid type) is obtained by mixing sophorolipid with alkali at room temperature.

  In one embodiment, the sophorolipid is substantially 100% acid sophorolipid.

  In one embodiment, 0.01 to 20% acid sophorolipid is included.

  In one embodiment, it further comprises a detergent supplement.

  In one embodiment, the detergent auxiliary component is at least one selected from the group consisting of enzymes, alkaline agents, water softeners (Ca supplements), foam regulators, preservatives, and solvents.

  According to the present invention, since it contains a large amount of chemically stable acid sophorolipid even in a liquid, it is possible to provide a cleaning composition containing biosurfactants for various uses. Moreover, it is suitable for the washing | cleaning process in which low foaming property is required, such as jet washing.

  Embodiments of the present invention will be specifically described below.

  Sophorolipid is typically obtained by culturing microorganisms. For example, Candida bombicola, C.I. apicola, C.I. petrophilum, C.I. Produced by Candida yeasts such as bogoriens. Sophorolipid accumulates in a large amount (100 to 150 g / L) in the medium when these Candida yeasts are cultured with a high concentration of sugar and an oily substrate simultaneously.

  Typically, the sophorolipid is separated from the above-mentioned microorganism culture solution by a method such as centrifugation, decantation, or ethyl acetate extraction, and further washed with hexane to obtain a brownish brown substance. Since sophorolipid has a higher specific gravity than water, it can be easily separated into the lower layer if the culture solution is allowed to stand, and the sophorolipid obtained using this is a hydrated substance with a concentration of about 50%. At the time of preparation, it is easy to handle because it has a lower viscosity than those separated and purified with an organic solvent such as hexane. Note that “%” used in this specification represents “% by weight” unless otherwise noted.

  The SL (sophorolipid) thus obtained has a basic structure composed of sophorose or sophorose partially hydroxylated with hydroxyl group and hydroxy fatty acid, and the acid type sophorolipid in which the carboxyl group of hydroxy fatty acid is liberated, It is a mixture of a plurality of molecular species roughly classified into a lactone type sophorolipid in which a carboxyl group is ester-bonded with a hydroxyl group of intramolecular sophorose.

  The acid-type sophorolipid contained in the low-foaming detergent composition described herein has, for example, a structure in which the lactone part of sophorolipid produced by microorganisms and separated and purified and the acetyl part of sophorose are all hydrolyzed. Again, it is a mixture of a plurality of structural analogs.

  The method for preparing the acid-type sophorolipid is not particularly limited, but a preparation method by alkali reflux is common. In this case, although depending on the subsequent purification method, acid-type sophorolipid can be obtained with a purity close to 100%. Alternatively, it can also be obtained by alkaline hydrolysis at room temperature as described herein.

The composition of the present invention contains at least 90% of sophorolipid (acid type) of sophorolipid. Preferably it is 95% or more, More preferably, it is 98% or more, Most preferably, it is acid form sophorolipid whose sophorolipid is substantially 100%.
The cleaning composition of the present invention preferably contains 0.01 to 20% of the acid type sophorolipid. When the content of the acid-type sophorolipid is less than 0.01%, effective surface activity cannot be obtained, so that the cleaning power is low. When the content exceeds 20%, the cleaning power decreases due to excessive foaming during cleaning. A more preferable content of acid sophorolipid is 0.1 to 5%.

  As used herein, the term “chemical stability” refers to the property of being blended as an alkaline cleaner above pH 10. Specifically, it means that the structure does not change in a sodium hydroxide solution having a pH of 13.7 for one month.

The term “low-foaming property” used in the present specification is a property exhibiting foaming power suitable for a washing process in which low-foaming property is required. Specifically, in the Ross Miles method, which is a commonly used method for evaluating foaming power, the foam height immediately after the flow is within 57 mm, and the foam after 5 minutes. The height is within 30mm. When the foam height exceeds 57 mm or 30 mm, problems such as a reduction in cleaning power due to a drop in jet water pressure due to foaming and foam overflow from the washing machine occur in cleaning using jet cleaning.
The term “excellent detergency” used in the present specification is a property exhibiting a cleaning performance equivalent to or higher than that of a conventional low-foaming surfactant suitable for a cleaning process requiring low-foaming property. Specifically, it is determined by performing a detergency test using a contaminated cloth, which is a method for evaluating detergency generally performed at present.

  The term “enzyme stability” as used herein is a property that exhibits enzyme stability equivalent to or higher than that of a conventional enzyme-containing liquid detergent. Specifically, the cleaning enzyme blended in the liquid detergent composition has the property that 80% or more of the enzyme activity remains after a storage stability test at 40 ° C. for one month.

The low-foaming detergent composition of the present invention may contain a cleaning auxiliary component in addition to the surfactant.
The cleaning auxiliary component may be the same as a known cleaning composition used in a jet cleaning system that requires low foaming properties, and is not particularly limited. For example, enzymes, alkaline agents, water softeners (Ca scavengers), rust preventives, foam modifiers and solvents that are currently formulated as dedicated detergent compositions for washer-disinfectors used in medical settings This is the case.

  The enzyme is one or more components selected from the group consisting of amylase, protease, cellulase, lipase, pullulanase, isopullulanase, isoamylase, catalase and peroxidase.

  The alkaline agent is at least one component selected from the group consisting of sodium hydroxide, potassium hydroxide, carbonic acid, hydrogen carbonate, silicic acid, metasilicic acid and alkali metal salts of boric acid, various ethanolamines, etc., and raises the pH. In this way, the cleaning power is strengthened, and when the enzyme is blended, the effect is also increased.

The Ca scavenger may be either an organic chelating agent or a polymer chelating agent.
The organic chelating agent is at least one component selected from nitrilotriacetate, ethylenediaminetetraacetate, citrate, succinate, polyphosphate, and the like.

  The polymer chelating agent is at least one component selected from a polymer of acrylic acid, methacrylic acid, maleic anhydride, α-hydroxyacrylic acid, itaconic acid, a copolymer thereof, and a salt thereof. . However, care must be taken in selecting the enzyme from the standpoint of enzyme stability.

  The solvent is used for enzyme stabilization and the like. It is selected from ethanol, glycerol, ethylene glycol, diethylene glycol, propylene glycol, hexylene glycol, polyethylene glycol and the like.

  The rust preventive is blended for the purpose of preventing corrosion of the washing machine and the object to be cleaned. It is selected from phosphate, chromate, nitrite, silicate, borate, carbonate, molybdate, benzotriazole, octadecylamine and the like.

  The content and type of the cleaning auxiliary component can be appropriately selected by those skilled in the art depending on the intended form and use of the cleaning composition.

  When preparing a low-foaming detergent composition, the content of the cleaning auxiliary component varies depending on the type of the component, but is selected to be 99.99% or less of the low-foaming detergent composition. Preferably it is 30 to 50%.

  The following examples illustrate the invention in more detail. The following examples are illustrative of the present invention and do not limit the present invention.

The evaluation items and test methods performed in the following examples are as follows.
1. Establish esterification rate evaluation system
In order to know how much the ester bond of SL was hydrolyzed, the ester value used for the analysis of fats and oils was used as an index. The ester value is expressed as the number of mg of KOH required to completely saponify (hydrolyze) the ester contained in 1 g of the sample. After neutralizing the sample, it was heated in a KOH ethanol solution to be saponified and consumed. The amount of KOH is determined from the decrease in neutralization equivalent. Therefore, if the initial neutralization equivalent is determined by titration at this time, the acid value (mg number of KOH required to neutralize 1 g of sample) and the saponification value can be known at the same time.

Ester decomposition rate (%) = (ester value before alkali treatment−ester value after alkali treatment) / ester value before alkali treatment × 100 (Formula 1)
In order to accurately determine the ester decomposition rate, it is necessary to know the ester value when the ester bond of SL before alkali treatment is completely hydrolyzed, so consider the heating time required for the reaction to proceed completely did.

  As samples, hydrous SL prototypes and purified samples from jar cultures were used. The production and storage environment of each sample, standard values, etc. are as shown in Tables 1 and 2 below.

エステル価の測定は、日本油化学会編 基準油脂分析法2.3.3-1996エステル価に従った。煮沸還流時間を０．５、１および２時間としてエステル価の変化を調べた。以下に操作手順を示す。
（１）含水SL〜2.5g（乾燥残分として〜1.5g）を中性エタノール50mlに溶解
（２）フェノールフタレインを指示薬とし、0.1 M KOHで中和滴定（→酸価）
（３）0.5 NKOH/EtOH 25 mlを正確に添加
（４）煮沸還流（0.5,1,2h）・冷却
（５）フェノールフタレインを指示薬とし、0.5N HClで中和滴定
２．アルカリ混合による室温下での酸型ソホロリピッド調製法
実施例1で求めたけん化価（表７）より、NaOH必要量はそれぞれ
ＳＬ−８Ｍ：239.3/1000×40.00/56.11＝0.170ｇ・ｇ^-1 （式４−ａ）
ＳＬ−２Ｍ：248.4/1000×40.00/56.11＝0.177ｇ・ｇ^-1 （式４−ｂ）
となる。

The ester value was measured according to the standard oil analysis method 2.3.3-1996 ester value edited by the Japan Oil Chemists' Society. Changes in the ester value were examined with boiling reflux times of 0.5, 1 and 2 hours. The operation procedure is shown below.
(1) Hydrous SL ~ 2.5g (~ 1.5g as dry residue) dissolved in 50ml of neutral ethanol (2) Neutralization titration with 0.1M KOH using phenolphthalein as indicator (→ acid value)
(3) Accurately add 25 ml of 0.5 NKOH / EtOH (4) Boiling reflux (0.5, 1, 2 h) and cooling (5) Neutralization titration with 0.5N HCl using phenolphthalein as an indicator Preparation method of acid type sophorolipid at room temperature by alkali mixing From the saponification value obtained in Example 1 (Table 7), the required amount of NaOH is SL-8M: 239.3 / 1000 × 40.00 / 56.11 = 0.170 g · g ⁻¹ ( Formula 4-a)
SL-2M: 248.4 / 1000 × 40.00 / 56.11 = 0.177 g · g ⁻¹ (Formula 4-b)
It becomes.

  Since it is considered that an excessive amount of alkali is necessary to proceed the ester decomposition reaction to near the end point, the amount of NaOH to be mixed was set to more than twice the necessary amount.

Two types of hydrous SL (SL-8M and SL-2M) were used as samples. An equal amount of NaOH aqueous solution containing NaOH twice or four times the amount required for decomposition was mixed with each hydrous SL, and allowed to stand at room temperature. After 30 minutes to 2 days, SL alkaline solution ˜5 g (˜1.5 g as SL dry residue) was collected, and after weighing, neutralized with about 2N HCl using phenolphthalein as an indicator. Hereinafter, the ester value was measured by the same method as Test Method 1. The boiling reflux time was 2 hours.
3. Sophorolipid Stability The acid type sophorolipid prepared as described in 2 above (reaction for 2 days) was stored at 50 ° C. for 1 month, and the stability was examined by comparing the HPLC chart immediately after the reaction and after storage.
4). HPLC analysis The lactone content of SL obtained by each method was determined by HPLC analysis. The conditions were a Nagel (Germany) Nucleosyl 5SB packed column (4.6 mm x 250 mm), 0.2% (w / v) sodium perchlorate / methanol solution as the mobile phase, column temperature 35 ° C, flow rate Detection was performed at an absorbance of 205 nm at 1 ml / min or with a differential refractometer.
5. Preparation of buffer 5-1. Menzel buffer
M / 50 sodium carbonate and sodium hydrogen carbonate were prepared and mixed at a ratio of 4.0: 6.0 to obtain a pH 8.94 (18 ° C.) buffer solution.
5-2. Preparation of hard water (AOAC method Synthetic Hard Water (AOAC official method of analysis (1995). Chapter 6, p9)
1 ml of 1 liquid was diluted with 60 ml or more of distilled water and 4 ml of 2 liquids were added to make up to 100 ml (hardness is about 1000 ppm). This was diluted with the above buffer solution to obtain hard water having the intended pH and hardness.
1st solution: 67.71 g of magnesium chloride hexahydrate and 73.99 g of calcium chloride were dissolved in distilled water and made up to 1 L.
Second solution: 56.03 g of sodium bicarbonate was dissolved in distilled water and made up to 1 L.
6). Foam test 6-1. Los Miles Act
Based on JIS K 3362, the foaming power and foam stability were measured using the RossMiles method. In this method, the above 5. A surfactant as a test substance was dissolved in the target concentration in the hard water prepared in the preparation of the buffer solution to obtain a test solution.
200 ml of each test solution was dropped on the liquid surface in 30 seconds from a height of 900 mm at a predetermined temperature condition (40 ° C. in this test), and the foam height immediately after that was used as the foaming power. The height after 5 minutes was defined as the foam stability.
6-2. Method using a dishwasher Various household dishwashers were used. Each detergent is put into a dishwasher so as to have a predetermined concentration (prototype WA, WA2, M-700L; 0.5%, M-251L; 1.0%), and foaming after 3, 5 and 10 minutes from the start of washing ( Immediately after opening the lid; 0 seconds later) and the speed of defoaming of the generated foam (5 seconds after the lid was opened) was visually observed and judged based on the foaming evaluation in Table 3.

７．洗浄力試験
７−１．ターゴトメーターを用いた洗浄力試験
ターゴトメーターは、栄科学精機製作所製、型式TM-4を使用した。試験カップに蒸留水または所定の硬度の人工硬水990 mLを入れ、30℃で保温した。各カップに表4の各試験液10 gをいれ、湿式人工汚染布(5cm×5 cm)(財団法人 洗濯科学協会）4枚を投入して120 rpmで20分間洗浄した。洗浄終了後、汚染布をピンセットで取り出し、1 Lの蒸留水で2回すすぎ、自然乾燥させた。洗浄率は、洗浄前後の反射率を色彩色差計(ミノルタ社製CR-300)により測定し、式４から洗浄率を算出した。
(式４) 洗浄率（％）＝(洗浄後の反射率−洗浄前の反射率)／(未汚染布の反射率−洗浄前の反射率)×100
７−２．食洗機を用いた洗浄力試験
食洗機を用い、５分間洗浄を行った。被洗浄物として、ステンレステストピースにレバー汚染液（牛生レバーに半重量の蒸留水を加えホモジナイズを行ったもの）を筆にて塗布し一晩乾燥させたものを用いた。洗浄開始時の温度は４０℃±１．５℃、洗浄終了時の温度は５０℃±１．５℃と設定した。洗浄力の評価は表に従い行った。

7). Detergency test 7-1. Detergency test using a targotometer A targotometer manufactured by Eikai Seiki Seisakusho, Model TM-4 was used. Distilled water or 990 mL of artificial hard water with a predetermined hardness was placed in a test cup and kept warm at 30 ° C. 10 g of each test solution shown in Table 4 was placed in each cup, and 4 wet artificially contaminated cloths (5 cm × 5 cm) (Laundry Science Association) were added and washed at 120 rpm for 20 minutes. After washing, the contaminated cloth was taken out with tweezers, rinsed twice with 1 L of distilled water, and allowed to air dry. For the cleaning rate, the reflectance before and after cleaning was measured with a color difference meter (CR-300 manufactured by Minolta), and the cleaning rate was calculated from Equation 4.
(Formula 4) Cleaning rate (%) = (Reflectance after cleaning−Reflectivity before cleaning) / (Reflectivity of uncontaminated cloth−Reflectivity before cleaning) × 100
7-2. Detergency test using a dishwasher A dishwasher was used for washing for 5 minutes. As an object to be cleaned, a stainless test piece was applied with a liver contamination solution (a homogenized product of half-distilled water added to cattle liver) and dried overnight. The temperature at the start of washing was set to 40 ° C. ± 1.5 ° C., and the temperature at the end of washing was set to 50 ° C. ± 1.5 ° C. Detergency was evaluated according to the table.

８．酸型ソホロリピッド配合液体洗浄剤組成物の調製
８−１．酸型ソホロリピッド配合アルカリ洗浄剤
洗浄機用のアルカリ洗剤として、表１に示す試作品を作製し、食洗機を用いた泡立ち試験および振盪洗浄力試験に供した。比較として、他社製品（M-700LおよびM-251L（いずれもシャープ製））を用い、同様に試験した。

8). 8. Preparation of acid type sophorolipid-containing liquid detergent composition 8-1. Acid Type Sophorolipid-Containing Alkali Detergent As an alkaline detergent for a washer, prototypes shown in Table 1 were prepared and subjected to a foaming test and a shaking detergency test using a dishwasher. As a comparison, other companies' products (M-700L and M-251L (both manufactured by Sharp)) were used and tested in the same manner.

  All prototypes were made by stirring with a stirrer. The caustic potash solution and 1K potassium silicate were added to the acid type SL solution prepared by an arbitrary method in this order and mixed. In the case of a pure soap formulation, it was added here, and finally distilled water was added. After confirming that the mixture was completely mixed visually, the following raw materials were added.

８−２．酸型ソホロリピッド配合中性酵素洗剤
各洗浄剤は、酸型ソホロリピッドに残りの原料を添加し調製した。このとき、完全に溶解したことを確認してから次の原料を添加した。

8-2. Acid type sophorolipid-containing neutral enzyme detergent Each detergent was prepared by adding the remaining raw materials to acid type sophorolipid. At this time, the next raw material was added after confirming complete dissolution.

９．プロテアーゼ活性の測定
カゼインを基質に用いたFolin-Lowry反応による測定を行った。
〔基質の調製〕
２％カゼイン溶液：２ｇのカゼイン（Hammerstein法で調製したもの）に0.1 N NaOH ２０mlを加え、加温しながら溶かす。室温まで冷却し、０．２MTris(hydroxymethyl) aminomethaneを１０ml加え、 HClで所定のpHに合わせ、水を加えて全量を100mlとする。
〔発色試薬〕
・ストック溶液A：１％CuSO₄・５H_２O,ストック溶液B：２%酒石酸カリウム, ストック溶液C：２％無水炭酸ナトリウム
・０．４M トリクロロ酢酸
０．４M Na_２CO_３
２ ｍｇ／ｍｌ チロシン溶液
〔活性測定〕
基質溶液０．５mlを３０℃に加温し、あらかじめ３０℃に保っておいた酵素液（洗浄剤組成物）０．５mlを加え、３０℃で１０分間インキュベーションした。氷冷した５％ TCAを１ｍｌ加え、氷中で約１０分間放置した後、遠心(15,000rpm、10 min、４℃)処理した。遠心上清に反応試薬（ストック溶液A,BおよびCを１：１：100で混合したもの）１．０ｍｌを加え、よく攪拌した後、室温で１０分間放置した。これにフェノール試薬を０．１ｍｌを加え、よく攪拌し、４０℃で１０分間放置後、直ちに氷冷した。分光光度計を用い660nmの波長で試料の吸光度を測定した。プロテアーゼ活性は、以下のように求めた。
プロテアーゼ活性＝（各試料の吸光度）―（各試料のブランクの吸光度）
（実施例１）エステル分解率評価系の確立
１．エステル分解率評価系の確立に従い、発酵で得られたソホロリピッドの加水分解を行った。図１に加熱還流時間と反応（けん化）（エステル分解）に必要としたKOH（ｍｇ／ｇ試料乾燥残分）の関係を、表７に各試料の酸価、２時間の反応に必要としたKOH（mg／g試料乾燥残分）およびけん化価を示す。

9. Measurement of protease activity Measurement was performed by Folin-Lowry reaction using casein as a substrate.
(Preparation of substrate)
2% casein solution: Add 20 ml of 0.1 N NaOH to 2 g of casein (prepared by Hammerstein method) and dissolve while warming. Cool to room temperature, add 10 ml of 0.2 M Tris (hydroxymethyl) aminomethane, adjust to a predetermined pH with HCl, and add water to make a total volume of 100 ml.
[Coloring reagent]
Stock solution A: 1% CuSO ₄ .5H ₂ O, stock solution B: 2% potassium tartrate, stock solution C: 2% anhydrous sodium carbonate, 0.4M trichloroacetic acid 0.4M Na ₂ CO ₃
2 mg / ml tyrosine solution [activity measurement]
0.5 ml of the substrate solution was heated to 30 ° C., 0.5 ml of an enzyme solution (cleaning composition) previously kept at 30 ° C. was added, and incubated at 30 ° C. for 10 minutes. 1 ml of ice-cooled 5% TCA was added, left in ice for about 10 minutes, and then centrifuged (15,000 rpm, 10 min, 4 ° C.). To the supernatant, 1.0 ml of a reaction reagent (stock solutions A, B and C mixed at 1: 1: 100) was added, stirred well, and allowed to stand at room temperature for 10 minutes. To this was added 0.1 ml of a phenol reagent, stirred well, allowed to stand at 40 ° C. for 10 minutes, and immediately cooled on ice. The absorbance of the sample was measured at a wavelength of 660 nm using a spectrophotometer. Protease activity was determined as follows.
Protease activity = (absorbance of each sample)-(absorbance of blank of each sample)
(Example 1) Establishment of ester decomposition rate evaluation system In accordance with the establishment of the ester degradation rate evaluation system, the sophorolipid obtained by fermentation was hydrolyzed. FIG. 1 shows the relationship between the heating reflux time and the KOH (mg / g sample dry residue) required for the reaction (saponification) (ester decomposition), and Table 7 shows the acid value of each sample required for the reaction for 2 hours. KOH (mg / g sample dry residue) and saponification value are shown.

２時間の反応に必要としたKOH（mg／g試料乾燥残分）は試料により異なり、加熱時間に関わらず常にSL−2MよりSL−８Mで大きかったが、０．５時間の加熱で急激に上昇し、以降は緩やかな増加に留まるという傾向は同じであった。以後の実験では煮沸還流を2時間とし、このときの値をエステル価とした（エステル分解率100％）。
（実施例２）アルカリ混合による室温下での酸型ソホロリピッド調製法
２．アルカリ混合による室温下での酸型ソホロリピッド調製法に従い、発酵で得られたソホロリピッドを室温下で加水分解を行った。表７に実験に使用した試料の乾燥残分当たりのエステル価（アルカリ処理前後）およびエステル分解率を、図２にアルカリ処理中のエステル分解率の経時変化をそれぞれ示した。

The KOH (mg / g sample dry residue) required for the reaction for 2 hours was different depending on the sample, and it was always larger at SL-8M than SL-2M regardless of the heating time, but it rapidly increased with heating for 0.5 hour. The trend was the same, rising and staying moderate thereafter. In subsequent experiments, boiling reflux was 2 hours, and the value at this time was defined as the ester value (ester decomposition rate 100%).
(Example 2) Preparation method of acid type sophorolipid at room temperature by alkali mixing According to the acid-type sophorolipid preparation method at room temperature by alkali mixing, the sophorolipid obtained by fermentation was hydrolyzed at room temperature. Table 7 shows the ester value (before and after alkali treatment) and the ester decomposition rate per dry residue of the sample used in the experiment, and FIG. 2 shows the change over time in the ester decomposition rate during the alkali treatment.

各含水SLに計算より求めた分解必要量の２倍のNaOHを添加し、約2時間以上静置することで９０％以上のエステル結合が分解された（図２）。また、放置時間を延ばしたり、NaOH添加量をさらに増やしてもエステル分解率は上昇しなかった（図２および表８）。これらの結果は含水SLの種類によらず同じであり、アルカリ処理後のエステル価も同程度であった（表８）。さらに、２時間の煮沸還流を行った試料と室温アルカリ処理した試料を、同条件でＨＰＬＣに供したところ、９０％以上が酸型SLであることが示された。以上から、短時間に比較的簡単な方法で大部分が酸型SLからなるSL組成物を水溶液として得られることが分かった。
（実施例３）酸型ソホロリピッドの起泡力
上記６−１．ロスマイルス法に従って、CaCO_３ ０から３００ppm、ｐH８．９４（１８℃）の条件下で所定の濃度の酸型ソホロリピッドの起泡力を調べた（図３）。その結果、酸型ソホロリピッド１％濃度では、硬度４０〜３００ｐｐｍまで、０．１％濃度では８〜２００ｐｐｍまで、０．０１％では０ｐｐｍであっても低泡性を示すことが判った。
（実施例５）酸型ソホロリピッドの洗浄力
７−１．ターゴトメーターを用いた洗浄力試験に従い、CaCO_３ ０から３００ppm、ｐH８．９４（１８℃）の条件下で１％濃度の酸型ソホロリピッドの洗浄力を調べた。図４から酸型ソホロリピッドは、水の硬度に関わらず、高い洗浄力を示すことが分かった。
（実施例７）酸型ソホロリピッドの安定性
３．酸型ソホロリピッドの安定性に準じて試験を行った（図５）。その結果、保存前後でHPLCチャートに変化はなく、酸型ソホロリピッドは優れた安定性を有していることが確認された。
（実施例６）酸型ソホロリピッド配合アルカリ洗浄剤の起泡性および洗浄力
表２の試作品について、６−２．食洗機を用いた方法に従い起泡性試験を行った（表９）。試験濃度は１．０％である。その結果、泡調整剤を配合しない場合、酸型ソホロリピッドは２０％配合で運転異常が生じたが、泡調整剤を配合することで、２０％配合が可能であった。

90% or more of the ester bond was decomposed by adding NaOH twice the amount required for decomposition to each hydrous SL and allowing to stand for about 2 hours or longer (FIG. 2). Further, the ester decomposition rate did not increase even when the standing time was extended or the amount of NaOH added was further increased (FIG. 2 and Table 8). These results were the same regardless of the type of hydrous SL, and the ester value after the alkali treatment was similar (Table 8). Furthermore, when a sample subjected to boiling reflux for 2 hours and a sample subjected to alkali treatment at room temperature were subjected to HPLC under the same conditions, 90% or more was shown to be acid type SL. From the above, it has been found that an SL composition consisting mostly of acid-type SL can be obtained as an aqueous solution in a relatively simple manner in a short time.
(Example 3) Foaming power of acid sophorolipid 6-1. According to the Ross Miles method, the foaming power of acid-type sophorolipid at a predetermined concentration was examined under the conditions of CaCO ₃₀ to 300 ppm and pH 8.94 (18 ° C.) (FIG. 3). As a result, it was found that even when the acid type sophorolipid concentration was 1%, the hardness was 40 to 300 ppm, the 0.1% concentration was 8 to 200 ppm, and 0.01% was 0 ppm even at 0 ppm.
(Example 5) Detergency of acid type sophorolipid 7-1. According to the detergency test using a tartometer, the detergency of 1% acid type sophorolipid was investigated under the conditions of CaCO ₃₀ to 300 ppm and pH 8.94 (18 ° C.). FIG. 4 shows that acid type sophorolipid exhibits a high detergency regardless of the hardness of water.
(Example 7) Stability of acid-type sophorolipid A test was conducted according to the stability of acid sophorolipid (FIG. 5). As a result, there was no change in the HPLC chart before and after storage, and it was confirmed that the acid-type sophorolipid has excellent stability.
Example 6 Foamability and Detergency of Acid Type Sophorolipid-Containing Alkali Detergent Regarding Prototypes in Table 2, 6-2. A foaming test was conducted according to a method using a dishwasher (Table 9). The test concentration is 1.0%. As a result, when no foam modifier was blended, 20% blending of acid type sophorolipid resulted in abnormal operation, but 20% blending was possible by blending the foam modifier.

次に、試作品Ａ１およびＡ２について、７−１．ターゴトメーターを用いた洗浄力試験を行った。結果を図６に示した。洗剤なしおよび他社商品M-251Lは比較対照である。この結果、酸型ソホロリピッド配合の２種の試作品は、いずれも洗浄率が８０％前後で、対照として用いたM-251Lと同程度の優れた洗浄力を有していた。
（実施例７）酸型ＳＬ配合＋洗浄補助成分＋酵素配合洗浄剤の洗浄力
試作品N1〜N8について、７−２．食洗機を用いた洗浄力試験を行った（図7）。その結果、酸型SL未配合のN1および酸型SLを２５％配合したN８の洗浄力は低く（ランクの平均が高い）、優れた洗浄力を示した配合はN2〜N7（０．０１〜２０％配合）であった。
（実施例８）酸型ＳＬ配合＋洗浄補助成分＋酵素配合洗浄剤の酵素保存安定性
試作品N２〜N６を４０℃のインキュベーター内に静置することで酵素安定性試験を行った。1週間毎にサンプリングを行い、８．プロテアーゼ活性の測定に従い測定し、保存前の酵素活性を１００％として酵素安定性を求めた（図８）。その結果、全ての試作品の酵素残存活性が８０％以上であり、酸型ソホロリピッド配合の洗浄剤が優れた酵素安定性を有していることが判明した。

Next, for prototypes A1 and A2, 7-1. A detergency test using a targotometer was performed. The results are shown in FIG. No detergent and other company product M-251L are comparative controls. As a result, the two types of prototypes containing the acid type sophorolipid each had a cleaning rate of around 80% and had excellent cleaning power comparable to that of M-251L used as a control.
Example 7 Detergency of Acid Type SL Formulation + Cleaning Auxiliary Component + Enzyme Formulation Detergent For Prototypes N1 to N8, 7-2. A detergency test using a dishwasher was performed (Fig. 7). As a result, N1 containing no acid type SL and N8 containing 25% acid type SL have low detergency (average rank is high), and N2 to N7 (0.01 to 0.01) 20% formulation).
(Example 8) Enzyme storage stability prototypes N2 to N6 of acid type SL formulation + cleaning auxiliary component + enzyme formulation detergent were subjected to enzyme stability test by standing in an incubator at 40 ° C. Sampling every week, 8. It was measured according to the measurement of protease activity, and enzyme stability before storage was determined as 100% to determine enzyme stability (FIG. 8). As a result, it was found that the residual enzyme activity of all prototypes was 80% or more, and the detergent containing the acid type sophorolipid had excellent enzyme stability.

The figure which shows the relationship between heating reflux time and KOH (mg / g sample dry residue) required for reaction (saponification). The figure which shows each time-dependent change of the ester decomposition rate during an alkali treatment. The figure which shows the influence (Upper: foaming power, Lower: Foam stability) which the hardness of water has on the foamability of acid-type sophorolipid. The figure which shows the influence which the hardness of water has on the detergency of acid-type sophorolipid. The figure which shows the storage stability test of acid type sophorolipid. A figure showing a comparison of the cleaning power of the prototypes WA2 and WA and the products of other companies. The figure which performed the detergency test using the dishwasher. The figure which measured according to the measurement of protease activity, and calculated | required enzyme stability by making the enzyme activity before storage into 100%.

Claims (6)

A biodegradable liquid detergent composition comprising sophorolipid, wherein the sophorolipid comprises at least 90% or more of sophorolipid (acid type). The composition according to claim 1, wherein the sophorolipid (acid type) is obtained by mixing sophorolipid with an alkali at room temperature. The composition according to claim 1, wherein the sophorolipid is substantially 100% acid-type sophorolipid. The composition according to any one of claims 1 to 3, comprising 0.01 to 20% of an acid-type sophorolipid. The composition according to claim 1, further comprising a detergent auxiliary component. The composition according to claim 5, wherein the detergent auxiliary component is at least one selected from the group consisting of an enzyme, an alkaline agent, a water softener (Ca supplement), a foam regulator, a preservative, and a solvent.

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