JP4072043B2 - Liquid enzyme for contact lenses - Google Patents

Description

【００３０】
表１から明らかなように、スルホブロモフタレインナトリウムを含有した酵素液は希釈液で希釈されてはじめて発色するため、使用者の注意をひくことが可能となっていることが確認された。
【００３１】
試験例１
希釈後の色の安定性試験
実施例１のスルホブロモフタレインナトリウム含有液体酵素剤を用いて希釈後の色の安定性を以下の方法で試験した。
【００３２】
１５ｍＬ容量のポリプロピレン製コニカルチューブにニューコンプリートを１０ｍＬ、分注した。ついで、実施例１の液体酵素剤を２００μＬ添加し、よく撹拌した。得られた混合液の吸光度（波長５８９．５ｎｍ）を分光光度計（（株）島津製作所製 ＵＶ−２４００ＰＣ）にて測定した。
【００３３】
つぎに、この混合液を２ｍＬずつコンタクトレンズ保存ケースに分注し、室温で放置した（ｎ＝５）。２４時間後にコンタクトレンズ保存ケースからポリプロピレン製コニカルチューブに混合液を回収し、吸光度を測定した。再度、混合液を２ｍＬずつコンタクトレンズ保存ケースに分注し、室温で放置した。前記混合調製から７日後に、２４時間後と同様にして混合液の吸光度を測定した。
【００３４】
以上で得られた結果を表２に示す。
【００３５】
【表２】

【００３６】
表２から明らかなように、表１の実施例１と希釈液における発色が混合時から少なくとも数日は持続し、使用者がコンタクトレンズを装用する時まで処理液が着色したままであることから、酵素処理を行なったコンタクトレンズをそのまま装用するといった誤用を防止することが可能であることが確認された。
【００３７】
試験例２
色素による固着タンパク質除去効果に対する影響（１）
本発明による液体酵素剤の固着タンパク質除去効果測定試験を以下の方法で実施した。
【００３８】
マンスウェア（（株）メニコン製 １ヶ月リプレースメントソフトコンタクトレンズ）をレンズケースに入れ、２ｍＬの２ｍｇ／ｍＬリゾチーム生理食塩水溶液を添加した。６０℃で約１時間、１００℃で約２０分熱処理し、そののち室温まで放冷することによりリゾチームの固着を行ないタンパク質固着コンタクトレンズを得た。
【００３９】
レンズを取り出し、生理食塩水で充分濯いだのち、生理食塩水を満たした１４．５φのセルにレンズを入れ、レンズの濁度（ＨＡＺＥ値）を積分球光電散乱光度計（日本電色工業株式会社製、ＮＤＨ−３００Ａ）を用いて測定した。測定は、セルを１２０°ずつ回転させ計３回行ない、その平均値をデータとした。なお、ＨＡＺＥ値０の対照として新品のマンスウェアを用いた。
【００４０】
得られたタンパク質固着コンタクトレンズを各処理液に浸漬し、室温で処理を開始させた。処理開始から４時間後にコンタクトレンズをとりだし、生理食塩水で充分濯いだあと、濁度を積分球光電散乱光度計を用いて測定した。処理液としては、実施例１の液体酵素剤４０μＬをアイネス（販売元：（株）メニコン マルチパーパスソリューション）２ｍＬで希釈したものを用い、対照として、酵素剤を含まないアイネスのみ、および色素を含まない比較例３の液体酵素剤４０μＬとアイネス２ｍＬとの混合液を使用した。各々の処理液に対して３枚のコンタクトレンズを使用した。
【００４１】
以下の計算式にしたがって、処理前および処理後の濁度の変化から各処理液の洗浄効果を計算した。結果を表３に示す。
洗浄効果（％）＝（処理前の濁度−処理後の濁度）／（処理前の濁度）×１００
【００４２】
【表３】

【００４３】
試験例３
色素による固着タンパク質除去効果に対する影響（２）
コンタクトレンズとしてメニコンフォーカス（販売元：（株）メニコン 頻回交換ソフトコンタクトレンズ）を用い、リゾチームの固着に２ｍＬの１ｍｇ／ｍＬリゾチーム生理食塩水溶液を用い、マルチパーパスソリューションとしてニューコンプリートを用いた以外は、試験例２と全く同様にして本発明の液体酵素剤の固着タンパク質除去効果測定試験を行なった。結果を表４に示す。
【００４４】
【表４】

【００４５】
表３および４から明らかなように、使用される色素（スルホブロモフタレインナトリウム）によって、洗浄効果が低下することはないことが確認された。
【００４６】
試験例４
蛋白質分解酵素の残存活性測定試験
本発明の液体酵素剤の酵素安定性を測定するために、液体酵素剤の調製から保存温度を４０℃として、１週間後、１ヵ月後および２ヶ月後の残存活性を以下の方法で測定した。
【００４７】
３７℃に加温した２．０％カゼイン溶液（ｐＨ７、０．０５Ｍ トリス−塩酸水溶液）４ｍＬに、希釈液（ｐＨ７、１０ｍＭ トリス−塩酸水溶液）の３．６ｍＬと、実施例１または比較例３の液体酵素剤０．４ｍＬをそれぞれ混合した。混和後ただちに、その１ｍＬを沈殿法（０．４ｍｏｌ トリクロロ酢酸水溶液による未分解カゼインの沈殿）にて、上澄み液中の２８０ｎｍにおける初期吸光度Ａ₀を求めた。残りの混合液を３７℃で１０分間保ち、ふたたび沈殿法にて、上澄み液中の２８０ｎｍにおける処理後吸光度Ａを求めた。処理後吸光度Ａより、初期吸光度Ａ₀を差し引くことにより、蛋白質分解酵素の活性値を数値化した。
【００４８】
残存活性（％）は次式により算出した。
酵素活性値［力価］＝Ａ−Ａ₀
残存活性値（％）＝［Ａ−Ａ₀］_DayX／［Ａ−Ａ₀］_Day0×１００
［Ａ−Ａ₀］_Day0：液体酵素剤調製日の酵素活性値
［Ａ−Ａ₀］_DayX：液体酵素剤調製日Ｘ日後（任意の期間後）の酵素活性値
結果を表５に示す。
【００４９】
【表５】

【００５０】
表５から明らかなように、使用される色素（スルホブロモフタレインナトリウム）によって、酵素安定性が低下することがないことが確認された。
【００５１】
また、試験例２〜４の結果から、本発明の液体酵素剤は色素を含有することによる影響もなく、コンタクトレンズに付着した汚れを充分に取り除くことができることが確認された。
【００５２】
【発明の効果】
本発明のコンタクトレンズ用液体酵素剤はマルチパーパスソリューションなどの希釈液に滴下した際に、はじめて処理液が発色することで、酵素処理していることを目で見て認識することができるため、使用者の注意を喚起し、酵素処理を行なったコンタクトレンズをそのまま使用するといった誤用を未然に防ぐことができる。
【００５３】
したがって、本発明の酵素液は、安全性、操作性に非常に優れた特徴をもっており、これによって使用者の目の安全性に多大な寄与を与えるものである。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a liquid enzyme agent for contact lenses. Specifically, the present invention relates to a liquid enzyme agent for contact lenses that develops color when diluted with a diluent such as a multipurpose solution.
[0002]
[Prior art]
Normally, liquid enzyme agents for contact lenses are colorless and transparent, and its use is known to drop the enzyme agent on the multi-purpose solution and immerse the contact lens, then rinse with the multi-purpose solution before wearing it. Yes. In addition, it is known that when the enzyme solution is not dropped and treated only with the multi-purpose solution, the lens is worn as it is without rinsing with tap water or physiological saline. Because of this method of use, the conventional colorless and transparent enzyme agent was treated by dropping the enzyme agent into a multi-purpose solution, but the user was inadvertently inadvertently removing the enzyme agent because it was colorless and transparent. It was misunderstood that it was not dripped, and it was easy to misuse that the lens was worn as it was without rinsing the lens treated with the enzyme. If an enzyme-treated lens is worn on the eyes without rinsing, eye damage such as inflammation may occur. Therefore, a system for eliminating such misuse is extremely useful in fields such as liquid enzyme agents for contact lenses, and its development has been strongly desired.
[0003]
As one of such misuse prevention systems, a method of coloring a diluent with a colorant is described (see Patent Document 1). In this method, the enzyme-containing cleaning solution is mixed with a preliminarily colored dedicated diluent, thereby facilitating identification of the cleaning solution and the storage solution prepared by mixing, and preventing misuse of the user. However, when such a diluted solution itself is colored in advance, there is a problem that it is difficult for a contact lens user to accept and the user's willingness to purchase is reduced.
[0004]
[Patent Document 1]
Japanese Patent No. 2624641 [0005]
[Problems to be solved by the invention]
In the present invention, in view of the prior art, the diluted solution and the liquid enzyme agent are colorless and transparent, and the liquid enzyme agent is diluted with the diluted solution and colored when used to alert the user and prevent misuse. It is an object to provide a system that enables the above.
[0006]
[Means for Solving the Problems]
The present invention finds that the liquid enzyme agent itself is not colored and can be prevented from being misused by using a liquid enzyme agent that contains a colorant that develops color when diluted. did.
[0007]
That is, the present invention relates to a liquid enzyme agent for a contact lens, which is a liquid enzyme agent containing a pigment, and the liquid enzyme agent itself is not colored but develops color when diluted with a diluent.
[0008]
In the liquid enzyme agent, the pigment is preferably a phenolphthalein pigment. As the phenolphthalein dye, sodium sulfobromophthalein is preferable.
[0009]
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, the present invention will be described in detail.
[0010]
Examples of the diluent in the present invention include a multipurpose solution. As used herein, “multipurpose solution” means a solution that can perform one or more of storage, washing, rinsing, and disinfection of contact lenses in a single solution, and many of these are commercially available (for example, Complete (Seller: AMO Japan Co., Ltd.).
[0011]
The coloring matter used in the present invention means a coloring matter for coloring a processing solution when dropped on a multipurpose solution or the like. Therefore, there is no particular limitation as long as it does not develop color in a liquid enzyme agent and develops color when diluted by adding it to a diluent such as a multi-purpose solution. However, since the pH of the multipurpose solution is generally near neutral, a phenolphthalein dye is preferable. Of these, sodium sulfobromophthalein, whose safety has been confirmed by the Japanese Pharmacopoeia, is more preferred. In addition, when sodium sulfobromophthalein is contained in a liquid enzyme agent, it does not develop color because it exists as a sodium salt. However, when the enzyme agent is mixed with a diluent, sodium is dissociated and visible light region. It has a remarkable feature that color development occurs by changing the absorption wavelength. Therefore, paying attention to such characteristics of sulfobromophthalein, it is considered to be very useful to use as a dye for a liquid enzyme agent that develops color when the enzyme agent is mixed with a diluent. In addition, the phenomenon of color development at the time of enzyme treatment has a strong visual impression, so it is considered effective for allowing the user to regularly practice enzyme treatment. Moreover, it is considered effective to reduce eye damage caused by lens contamination caused by forgetting enzyme treatment as in the past by making the treatment with an enzyme agent customary.
[0012]
The blending amount of the dye in the liquid enzyme agent is preferably 0.001 to 10 w / w%, and more preferably 0.01 to 1 w / w%. When the blending amount of the dye is less than 0.001 w / w%, there is a tendency that the user hardly recognizes because the color is hardly formed, and when it exceeds 10 w / w%, the action of the enzyme agent tends to be inhibited.
[0013]
The liquid enzyme agent in the present invention is not only a well-known liquid enzyme agent containing a polyhydric alcohol, a boric acid compound and a proteolytic enzyme, but also mixed with a diluent such as a multi-purpose solution. The liquid enzyme agent is not particularly limited.
[0014]
Enzymes contained in the liquid enzyme agent of the present invention include those that degrade proteins that are the main components of contact lens dirt, and those that degrade other dirt components. Specific examples include proteolytic enzymes, lipolytic enzymes, glycolytic enzymes, and the like, but are not limited to these types.
[0015]
The liquid enzyme agent for contact lenses of the present invention may further contain known auxiliaries such as chelating agents, preservatives, bactericides, pH adjusting agents (buffering agents) and / or surfactants. These auxiliaries allow the enzyme to be stably present in the liquid enzyme agent for contact lenses of the present invention or the final preparation of the enzyme agent, and adversely affect the target, human body and environment to which the enzyme agent of the present invention is applied. There is no particular limitation as long as it does not. Moreover, the addition amount of each adjuvant is not limited to the following illustrations, It can set suitably by those skilled in the art.
[0016]
Examples of the chelating agent include ethylenediaminetetraacetic acid and salts thereof, such as ethylenediaminetetraacetic acid / disodium and ethylenediaminetetraacetic acid / trisodium. The addition amount of the chelating agent is preferably 0.01 to 2 w / w%.
[0017]
Examples of the preservatives include mercury preservatives such as phenylmercuric nitrate, phenylmercuric acetate and thimerosal, surfactant preservatives such as benzalkonium chloride and pyridinium bromide, and piguanides such as chlorhexidine and polyhexamethylene biguanide. Alcohol-based preservatives such as chlorobutanol, methylparaben, propylparaben, dimethyloldimethylhydantoin, imidazolium urea, boric acid, boric acid compounds, or borax can be used. Among them, preferred is one or two or more substances selected from boric acid, boric acid compounds and borax, and the addition amount is 0.00001 to 4.0 w / w%. Preferably there is.
[0018]
As the bactericidal agent, an organic nitrogen type bactericidal agent is preferable since it has a low bactericidal effect at a low concentration.
[0019]
Representative examples of the organic nitrogen-based disinfectant include (1) quaternary ammonium compound or a polymer thereof, such as benzalkonium chloride, benzethonium chloride, and dimer 136, and (2) a biguanide compound or a polymer thereof or the like. Examples thereof include chlorhexidine gluconate and polyhexamethylene biguanide which are salts, and (3) polymers of the above (1) and (2). Among them, a biguanide compound or a derivative thereof (polymer or salt) is more preferable because of its high safety, and polyhexamethylene biguanide (PHMB) is very particularly preferable because it does not absorb or adsorb to a lens. The amount of the organic nitrogen-based fungicide added is about 0.000001 to 10 w / w%, preferably about 0.00001 to 1 w / w%.
[0020]
Examples of the pH adjusting agent (buffering agent) include boric acid and its salt, phosphoric acid and its salt, citric acid and its salt, lactic acid and its salt, amino acid such as glycine or glutamine and its salt, or malic acid The salt etc. can be used. The addition amount of the pH adjuster is preferably 0.001 to 4.0 w / w%.
[0021]
As the surfactant, for example, any surfactant such as an anionic surfactant, a nonionic surfactant, or an anionic surfactant and a nonionic surfactant may be used.
[0022]
Examples of the anionic surfactant include sodium alkyl sulfate, sodium alkylbenzene sulfonate, sodium alkyloylmethyl taurate, sodium alkyloyl sarcosine, sodium α-olefin sulfonate, sodium polyoxyethylene alkyl ether phosphate, polyoxyethylene alkyl Sodium ether sulfate, polyoxyethylene alkylphenyl ether sodium sulfate, di (polyoxyethylene alkyl ether) sodium phosphate, and the like can be used. The amount of the anionic surfactant added is, for example, 0.01 w / v% or more, preferably 0.02 w / v% or more, and 10 w / v% or less, preferably 5 w / v% or less.
[0023]
Nonionic surfactants include, for example, polyethylene glycol adducts of higher alkylamines, polyethylene glycol adducts of higher fatty acid amides, polyglycerol esters of higher fatty acids, polyethylene glycol esters of higher fatty acids, polyalkylene glycols of higher fatty acids, polyethylene Glycol copolymer ester, polyhydric alcohol ester with higher fatty acid polyethylene glycol added, polyethylene glycol ether of higher alcohol, polyglycerin ether of higher alcohol, polyethylene glycol ether of alkylphenol, formaldehyde condensate of polyethylene glycol ether of alkylene phenol, polypropylene glycol -Polyethylene glycol copolymer, phosphate ester, castor oil Hydrogenated castor oil, polyethylene glycol sorbitan alkyl esters, polyethylene glycol adducts of sterols like can be used. The addition amount of the nonionic surfactant is, for example, 0.01 w / v% or more, preferably 0.02 w / v% or more, and 10 w / v% or less, preferably 5 w / v% or less.
[0024]
When the anionic surfactant and the nonionic surfactant are used in combination, the content of the anionic surfactant and the nonionic surfactant is within the above-described range, and the total amount is 0.02 to 20 w / v%, preferably 0.05 to 10 w / v%.
[0025]
【Example】
Hereinafter, the method of the present invention will be described in more detail with reference to examples. The following examples are for illustrative purposes only and are not intended to limit conditions and technical scope.
[0026]
Example 1 and Comparative Examples 1-3
<Preparation of liquid enzyme agent>
Prepare liquid enzyme agent at the ratio of glycerin 32w / w%, polyethylene glycol # 1000 28w / w%, boric acid 2w / w%, proteolytic enzyme (CL-5PG: manufactured by Nagase ChemteX Corp.) 3w / w% did.
[0027]
To the prepared liquid enzyme agent, sodium sulfobromophthalein sodium, phenol sulfonephthalein, and indicocarmine were added at a rate of 0.1 w / w%, respectively, to obtain Example 1 and Comparative Examples 1 and 2. Moreover, the liquid enzyme agent which has not added the pigment | dye was made into the comparative example 3.
[0028]
<Color inspection>
Using Example 1 and Comparative Examples 1 to 3, it was diluted 50 times with New Complete. The change in color before and after dilution was observed with the naked eye. The results are shown in Table 1.
[0029]
[Table 1]

[0030]
As is apparent from Table 1, since the enzyme solution containing sodium sulfobromophthalein was colored only after being diluted with the diluent, it was confirmed that the user's attention could be drawn.
[0031]
Test example 1
Color stability test after dilution Using the sodium sulfobromophthalein-containing liquid enzyme agent of Example 1, the color stability after dilution was tested by the following method.
[0032]
10 mL of New Complete was dispensed into a 15 mL polypropylene conical tube. Next, 200 μL of the liquid enzyme agent of Example 1 was added and stirred well. The absorbance (wavelength: 589.5 nm) of the obtained mixed solution was measured with a spectrophotometer (UV-2400PC manufactured by Shimadzu Corporation).
[0033]
Next, 2 mL of this mixed solution was dispensed into a contact lens storage case and allowed to stand at room temperature (n = 5). After 24 hours, the mixed solution was collected from the contact lens storage case into a polypropylene conical tube, and the absorbance was measured. Again, 2 mL of the mixture was dispensed into a contact lens storage case and allowed to stand at room temperature. Seven days after the preparation of the mixture, the absorbance of the mixture was measured in the same manner as 24 hours later.
[0034]
The results obtained above are shown in Table 2.
[0035]
[Table 2]

[0036]
As is apparent from Table 2, the color development in Example 1 and the diluent in Table 1 persists for at least several days from the time of mixing, and the treatment liquid remains colored until the user wears the contact lens. It has been confirmed that it is possible to prevent misuse of wearing contact lenses that have been subjected to enzyme treatment as they are.
[0037]
Test example 2
Effect on pigment removal effect by dye (1)
The fixed protein removal effect measurement test of the liquid enzyme agent according to the present invention was carried out by the following method.
[0038]
Manswear (1 month replacement soft contact lens manufactured by Menicon Co., Ltd.) was placed in the lens case, and 2 mL of 2 mg / mL lysozyme saline solution was added. After heat treatment at 60 ° C. for about 1 hour and at 100 ° C. for about 20 minutes, the mixture was allowed to cool to room temperature, whereby lysozyme was fixed to obtain a protein-fixed contact lens.
[0039]
After taking out the lens and rinsing it thoroughly with physiological saline, the lens is put into a 14.5φ cell filled with physiological saline, and the turbidity (HAZE value) of the lens is measured using an integrating sphere photoelectric scattering photometer (Nippon Denshoku Industries Co., Ltd.). Measurement was performed using NDH-300A manufactured by Co., Ltd. The measurement was performed three times by rotating the cell by 120 °, and the average value was used as data. As a control with a HAZE value of 0, a new monthwear was used.
[0040]
The obtained protein-fixed contact lens was immersed in each treatment solution, and treatment was started at room temperature. After 4 hours from the start of the treatment, the contact lens was taken out, rinsed thoroughly with physiological saline, and turbidity was measured using an integrating sphere photoelectric scattering photometer. As the treatment liquid, 40 μL of the liquid enzyme agent of Example 1 diluted with 2 mL of Ines (sales company: Menicon Multipurpose Solution) was used, and as a control, only Ines without the enzyme agent and the dye were included. A liquid mixture of 40 μL of the liquid enzyme agent of Comparative Example 3 and 2 mL of Ines was used. Three contact lenses were used for each treatment solution.
[0041]
According to the following calculation formula, the cleaning effect of each processing solution was calculated from the change in turbidity before and after the treatment. The results are shown in Table 3.
Cleaning effect (%) = (turbidity before treatment−turbidity after treatment) / (turbidity before treatment) × 100
[0042]
[Table 3]

[0043]
Test example 3
Effect on pigment removal effect by dye (2)
Except for using Menicon Focus (sales company: Menicon Co., Ltd., frequent exchange soft contact lens) as a contact lens, using 2 mL of 1 mg / mL lysozyme saline solution for fixing lysozyme, and using New Complete as a multi-purpose solution. In the same manner as in Test Example 2, a test for removing the fixed protein of the liquid enzyme agent of the present invention was conducted. The results are shown in Table 4.
[0044]
[Table 4]

[0045]
As apparent from Tables 3 and 4, it was confirmed that the washing effect was not lowered by the dye used (sulfobromophthalein sodium).
[0046]
Test example 4
Proteolytic enzyme residual activity measurement test In order to measure the enzyme stability of the liquid enzyme agent of the present invention, the storage temperature after preparation of the liquid enzyme agent is 40 ° C., and the residual after 1 week, 1 month and 2 months The activity was measured by the following method.
[0047]
To 4 mL of 2.0% casein solution (pH 7, 0.05 M Tris-hydrochloric acid aqueous solution) heated to 37 ° C., 3.6 mL of a diluent (pH 7, 10 mM Tris-hydrochloric acid aqueous solution) and Example 1 or Comparative Example 3 Each of 0.4 mL of the liquid enzyme agent was mixed. Immediately after mixing, 1 mL of the mixture was subjected to a precipitation method (precipitation of undegraded casein with a 0.4 mol trichloroacetic acid aqueous solution) to determine an initial absorbance A ₀ at 280 nm in the supernatant. The remaining mixed solution was kept at 37 ° C. for 10 minutes, and the post-treatment absorbance A at 280 nm in the supernatant was determined again by precipitation. By subtracting the initial absorbance A ₀ from the absorbance A after the treatment, the activity value of the proteolytic enzyme was quantified.
[0048]
The residual activity (%) was calculated by the following formula.
Enzyme activity [potency] = A-A ₀
Residual activity value (%) = [A−A ₀ ] _DayX / [A−A ₀ ] _{Day 0} × 100
[A-A _{0] Day0:} enzyme activity of the liquid enzyme preparation date of preparation [A-A _{0] DayX:} Liquid enzyme preparations prepared date X days after enzyme activity (after any period of time) Table 5 shows the results.
[0049]
[Table 5]

[0050]
As apparent from Table 5, it was confirmed that the enzyme stability was not lowered by the dye used (sulfobromophthalein sodium).
[0051]
In addition, from the results of Test Examples 2 to 4, it was confirmed that the liquid enzyme agent of the present invention can sufficiently remove the dirt adhered to the contact lens without being affected by containing a pigment.
[0052]
【The invention's effect】
When the liquid enzyme agent for contact lenses of the present invention is dropped into a diluent such as a multi-purpose solution, the treatment liquid is colored for the first time, so that the enzyme treatment can be visually recognized. It is possible to prevent misuse of alerting the user and using the contact lens that has been subjected to the enzyme treatment as it is.
[0053]
Therefore, the enzyme solution of the present invention has very excellent characteristics in safety and operability, and thereby greatly contributes to the safety of the eyes of the user.

Claims (1)

A liquid enzyme agent for contact lenses, which is a liquid enzyme agent containing sodium sulfobromophthalein as a coloring matter, and the liquid enzyme agent itself is not colored but develops color when diluted with a diluent.