JPS6227466A - Coloring compound - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to colored cationic compounds, especially those with cellulose-reactive groups, which are useful for protein adsorbents and protein adsorbents made from cationic compounds, which are important in the separation and purification of proteins. Regarding precipitating agents.

Prior Art For example, U.S. Pat. No. 4,016,149 and PCT No. 79
No. 00541 discloses that carbohydrate-based adducts such as polymers and copolymers derived from agarose, dextrose, dextran, etc., and certain commercially available cellulose-reactive acid dyes are used as adsorbents for chromatographic separation of protein materials. It is known that

DISCLOSURE OF THE INVENTION Novel colored compounds containing cellulose-reactive groups which may be attached to a substrate are disclosed and are suitable for the separation of biologically complex molecules that cannot be conveniently separated by known materials. Provides a highly absorbent adsorbent.

According to the present invention, a colored compound having a cationic group and a cellulose-reactive group is obtained.

Therefore, the charge may be bonded to the nitrogen atom, or may be delocalized over all cation groups or over all colored compounds. Advantageously, the cationic group is stable to changes in hydrogen ion concentration up to -11, especially up to pH 9.

Examples of suitable cationic groups are fully substituted ammonium groups, such as quaternary ammonium groups and their cycloaliphatic and aromatic analogues, and especially amisonium and guanidium groups. The cationic group may be a simple group such as a tetraalkylammonium group or an unsubstituted guanidinium or amisonium group, or it may have substituents.

The cellulose-reactive groups (hereinafter referred to as "reactive groups") are preferably mono- or borino-orinoheterocycles, such as dichloropyrimidinyl, difluoropyrimidinyl, trichloropyrimidinyl, 5-chlorodifluoropyrimidinyl. and 5-cyanodichloropyrimidinyl.Particularly preferred reactive groups are dichlorotriazinyl and monochlorotriazinyl groups.

The colored compound consists of a chromophore, preferably an azo chromophore and more preferably a monoazo chromophore, preferably attached to a reactive group, which chromophore has or incorporates a cationic group.

The colored compound is preferably of the formula: % wherein A represents a cationic group, Y represents a chromophore, Y' is a cationic chromophore and B is a reactive group. .

It has been found that colored compounds are advantageous over colorless compounds because colored compounds provide a direct visual indication of the degree of reaction with the carbohydrate substrate used in the production of protein precipitants or adsorbents.

Preferred cationic groups are: in which R1, R2, R3 and R4 represent optionally substituted alkyl, alkenyl or aryl groups, B represents an aromatic system, D represents an alicyclic system, and at least one of R1, R2, R3, R4, B and D has a free target; the cationic group itself is a non-chromophore; Often, the cationic group may then be pendant to the chromophore Y, or the cationic group may form part of the cationic chromophore Y'; if one or more of R1-R4 is attached to the chromophore For example, the group shown in formula (2) corresponds to AY- in formula I. RI M+ If R4 or two of B or D form part of a chromophore, the group shown in 2 corresponds to Y'- in formula 2; (i1) Formula: [wherein R3 , Rγ, R8 and R9 represent H or an optionally substituted alkyl, alkenyl or aryl group, and n represents 0 or 1]; the cationic group itself may be a non-chromophore. , in which case the di or cationic group pendant to the chromophore Y may constitute part of the cationic chromophore Y'. If the cationic group is not a chromophore, the cationic group is -(N
H) Advantageously bound to the chromophore Y via a free target of the group. If R6 and R7 or R8 and R9 form part of a chromophore, the group shown in formula
corresponds to When any one of R1++ R9 is alkyl or alkenyl, C1-4-alkyl or 0
Preference is given to 1-4-alkenyl, especially methyl or ethyl. R1% If any of R' is aryl, this is preferably a phenyl group cR1-R4
It is particularly advantageous that each of R6-R9 is H or an alkyl group.

The chromophore is preferably an azo chromophore, and preferred azo chromophores are derived from aniline or substituted aniline diazo moieties and from substituted benzene, naphthalene or pyrazolone coupling moieties, where the substituents are preferably C□ to 4- alkyl,
C1-4-alkoxy, NH2, NH2CO-1N) (
2CONH-, phenyl, 01-4-alkyl-Co
-C1-4-alkyl-CONH-101-4-alkyl-CONH-1OH and halogens, especially chlorine.

Examples of the group A-Y in formula (1) are: 4-(6- and 4-trimethylammoniophenylazo)-aniline.

4-(3- and 4-guanidinophenylazo)-6-methylphenylamino.

4- (5'- and 4'-amidinophenylazo)phenylamino.

4-<5- and 4-amidinophenylazo)-6-methylphenylamino.

4-(3- and 4-amidinophenylazo)-2,5-
Dimethylphenylamino.

4-(6-and 4-amidinophenylazo)-5-methyl/-2-methoxyphenylamino.

4-(6- and 4-amidinophenylazo)-22,5
-dimethoxyphenylamino.

4-(6-and 4-amidinophenylazo)-acetylaminoaniline.

4-(6- and 4-amidinophenylazo)-6-ureitophenylamine.

4-(6-and 4-amidinophenylazo)-6-nodoxy-6-acetylaminophenylamino.

4-(3- and 4-amidinophenylazo)-N-methylphenylamino.

4-((4-guanidinophenylazo)-(7cetyl)
-acetylamino)-phenylamino.

4-[: 4-(4-toIJ methylammoniophenylazo)-6-methylpyrazol-5-one-1-
Irkuphenylamino.

4-C4-(4-amidinophenylazo)-3-methylpyrazol-5-one-1-ylcuphenylamino,
and 4-C4-<4-guanidinophenylazo)-6-methylpyrazol-5-one-1-ylcuphenylamino.

Examples of suitable fully substituted ammonium groups are tetraalkylammonium groups, such as tetramethylammonium, aryl- and alkylkyl-trallykylammonium groups such as phenyltrimethylammonium and benzyltrimethylammonium, and especially when the phenyl group is azo Groups that are part of a chromophore, e.g. 4-(6-methysypheny-4-yl-azo)-phenyltrimethylammonium; cycloaliphatic ammonium groups, e.g. N,N-dimethylpyrrolidinium; cycloaliphatic ammonium groups, e.g. N
, N-dimethylpyrrolidinium; and mono- or polycyclic arylammonium groups, such as N-methylpyridinium and N-methylisoquinolinium, including those in which the heteroaromatic ring is part of the azo chromophore. be. Examples of suitable guanidinium and amidinium groups include guanidinium, amidinium and R6-R9 each independently H or 0.
The mono-tetra-alkyl-guanidinium and -amidinium groups shown above in Iv which are 1-4-alkyl, but advantageously all of R6-R9 are H. Particularly preferred guanidinium and amisonium groups are arylguanidinium and arylamizonium groups, ie, those in which the (NH)n group in (1) above is bonded directly to the azo chromophore. An example of such a group is 4-(
6-Methyl-4-[4-N,N,N'-trimethylguanizonoN'-phenylazo]-pyrazolin-5-one-1-yl)-phenyl and 4-(4-amidinophenylazo)- It is 2-methysyphenyl-1-yl.

The chromophore is advantageously attached to the reactive group via a linking group, for example an amino group -NR-, which linking group is advantageously derived from an amino group present on one of the benzene rings of the chromophore. Ru. Optionally, the linking group has the formula NR-X-NR- or -X-NR-, where X represents an optionally substituted divalent hydrocarbon, such as an optionally substituted alkylene or arylene; R may also be H1 alkyl, preferably C1-4-alkyl or aryl, preferably phenyl. Examples of such linking groups are alkylene diamines, such as ethylene diamino and arylene diamino, such as phenylene diamino and biphenylene diamino, aminoalkylenes, such as amine ethylene and aminopropylene, aminoarylenes, such as aminophenylene.

Particularly preferred reactive groups are of the formula is a mono- or dichlorotriazine group which is advantageously attached to the heterocycle. second cationic group A'
may be the same as or different from A and is advantageously bonded to the heterocycle via a linking group as described above. Examples of suitable non-reactive groups identified by T are mono- or bicyclic aryl, in particular phenyl, which may be H1-substituted, or an alkyl group. Examples of suitable groups represented by T-0- are optionally substituted alkoxy, such as methoxy,
Ethoxy, 1-propoxy, butoxy and ethoxyethoxy, and optionally substituted aryloxy, such as phenoxy, 4-methoxyphenoxy and naphtho-2-
It's oxy. Examples of suitable groups represented by T-NR- are NR2, methylamino, dimethylamino, ethylamine, 2-pyridinoethylamino, jetanolamino, anilino, 4-methylanilino, N-methylanilino, 6- or 4-trimethyl ammonioanilino, piperidino and morpholino.

Compounds of formulas I and II are preferably of the formula: A-Y-HVI, in which A and Y are as defined above and H is a polyhaloke9) hydrogen capable of reacting with the active halogen atom in the heterocyclic compound. It is advantageously produced by reacting the compound under conditions such that one atom remains.

An example of a suitable compound of formula (1) is 4-(3'-and 4'-trimethylammoniophenylazo)aniline chloride.

4- (3'- and 4'-guanidinophenyla f)-
6-Methylaniline chloride.

4-(3'- and 4'-amidinophenylazo)aniline chloride.

4- (3'- and 4'-amidinophenylazo)-6
-Methylaniline chloride.

4-(6'- and 4'-amidinophenylazo)-2,
5-dimethylaniline chloride.

4- (6'- and 4'-amidinophenylazo)-5
-Methyl-2-methoxyaniline chloride.

4- (3'- and 4'-amidinophenylazo)-2
, 5-dimethoxyaniline chloride.

4- (3'- and 4'-amidinophenylazo)-6
-acetylaminoaniline chloride.

4- (3'- and 4'-7midinophenylazo)-6
-Urateaniline chloride.

4- (3'- and 4'-amidinophenylazo)-6
-Nodoxy-6-acetylaminoaniline Chlori P.

4- (3'- and 4'-amidinophenylazo)-N
-Methylaniline chloride r.

4-([4-guanidino-phenylazo]-I[acetocouacetylamino)aniline chloride°.

1-(4-)limethyl-ammoniophenyl)-4-
(4-Aminophenylazo)-6-methyl-5-pyrazolone chloride.

1-(4-aminophenylazo)-4-(4-guanidinophenylazo)-6-methyl-5-pyrazolone chloride.

The compounds of formulas I and 1 have a particularly good affinity for proteins and are thus suitable for the production of protein adsorbents and precipitants as described above.

According to a second aspect of the invention, there is also provided an adsorbent consisting of an adduct of a cationic compound of formula (1) or formula (2) with a substrate, which may be a solid support or a soluble polymer. Examples of solid supports include acrylic polymers and copolymers, silica, titania, alumina, hydroalkyl methacrylates, porous glasses, but preferred solid supports are carbohydrate substrates. The carbohydrate substrate may be any solid carbohydrate material capable of reacting with reactive groups in a colored cationic compound, where the colored reactive compound is a carbohydrate, such as a polymer of 7-gallose, dextrose, dextran, and polyamide, especially polyacrylamide. and may be covalently bonded to the copolymer.

A particularly preferred carbohydrate is a polymer or copolymer of agarose.

The i-trix consisting of a solid support to which a dye is attached may be in the form of a column for chromatographic separation purposes or in the form of a membrane for separation carried out in a membrane separation mode.

Examples of soluble polymers reactive with colored cationic compounds include polyethylene glycol or dextran, the products of this reaction being used to separate protein materials in an aqueous two-layer partitioning process.

The adsorbent disclosed according to the second aspect of the invention can be prepared in accordance with standard methods, e.g. in the presence of an acid binder, e.g. an alkali metal carbonate or alkali metal hydroxide, such as sodium carbonate or sodium hydroxide. It is produced by reacting a compound with a carbohydrate substrate. The preparation of such adsorbents and chromatographic columns containing them is well documented (e.g. PCR K 79
00541 and US Pat. No. 4,016,149).

A sixth feature of the present invention is to obtain a protein precipitant comprising an adduct of two or more molecules of a colored cationic compound bonded via a polyfunctional bonding group. Such adducts can be used to precipitate proteins by aggregation, where they act as bridges between protein molecules. The adduct is composed of two or more molecules of a colored cationic compound and two or more molecules capable of reacting with a reactive group in the colored cationic compound.
It can be formed by reacting with a polyfunctional binder having more than one group. Examples of difunctional binding compounds are alkylene diamines such as tetramethylene diamine and hexamethylene diamine which may be condensable with colored cationic compounds. If the reactive group is a halogenoheterocycle, the reaction occurs with elimination of hydrogen halide. Examples of binding compounds with higher functionality are polyaminoalkanes such as diethylenetriamine and tetraethylenepentamine, which may be combined with more than 2 moles of colored cationic compounds.

Said adsorbents and precipitants derived from the compounds of the first aspect of the invention have a significantly enhanced binding capacity for certain biological molecules, such as proteins and enzymes, in particular proteases; It is especially applied to separate such types from mixtures of proteins.

Examples of enzymes and proteins to which the adsorbents and precipitants according to the invention are particularly adapted for binding are trypsin, thrombin and carboxypeptidase-B. They are therefore easily separated from the mixture of enzymes and proteins in which they are present.Examples The present application is illustrated in detail with reference to the following examples. Parts and percentages in the examples are shown in bold unless otherwise specified.

Example 1 4-Aminobenzamidine dihydrochloride in 400 g of water and 35 g of 2N hydrochloric acid 10.4. ! The solution of i' was cooled to below 5° C., and 25 g of 2N sodium nitrite solution was added. The solution was stirred for 15 minutes and the slight excess of nitrous acid was removed by adding a few drops of sulfamic acid bath. Then 6 g of 6-methylaniline was added to the bath solution, sodium acetate was added to bring the value to 4.5, and the mixture was stirred overnight. The resulting 4-(4=amino-2-methylphenylazo)benzamidine hydrochloride was separated off and dried.

4-(4) in a mixture of 350 g of water and 650 g of acetone
-Amino-2-methylphenylazo)benzamidine hydrochloride 7.61 of cyanuric chloride 6 in 100 g of water
．． 2 g of the water-cooled suspension, keeping the temperature of the mixture below 5 °C and carbonic acid at - value) Add as much of the 2N solution of IJum as desired to keep it at -6 to 7. .

Then, 4.8 I of 4-aminophenyltrimethylammonium chloride was added to the mixture, and the whole mixture was kept at pH 6-7 by adding a 2N solution of sodium carbonate.
The mixture was stirred at 60°C for 4 hours. Mainly 4-CA-C4,6-
dichloro-8-triazin-2-ylaminoco-2-methyl-phenylazo)benzamidinium chloride and a small amount of 4-(4-(4-10rho6-C4-trimethylammonioanilino)-8-triazine-2 A precipitate consisting of -ylaminoco-2-methyl-phenylazo)-benzamidinium dihydrochloride was formed, which was filtered off and dried.

Example 2 4-(4'-Amino-2'-methyl-
Phenyl 7yP) Benzamidinium Chloride 5.8
,! Add 6.8 g of cyanuric chloride to the ice-cold solution of i', -
The value was kept between 5 and 6 and finally 20.9 of 1M sodium hydroxide solution was added to bring the pH to 7. To this solution, 8 g of mixed phosphate buffer (equimolar amount of KH2PO4 and Na2HPO4) was added, followed by 40 g of salt, and the precipitate of dichlorotriazine was separated by salt filtration. 4-(4-(4,6
-dichloro-s-triacya2-i7mi)]-]2
-Methyl-phenylazopenzazonicum chloride was obtained.

Example 6 Water 500. ! Add 6.8 I of cyanuric chloride to an ice-cold solution of 4-aminophenyltrimethylammonium chloride 3.75 and F in i', and adjust the -value to 1 M NaOH solution 20
g was added to keep it at 5-6, but it rose to 7 in some cases. To the mixture was added 5.81 liters of 4-(4'-amino-2'-methylphenylazo)benzamidinium chloride, and the suspension was stirred at 40°C for 6 hours, during which 1M
By adding NaOH solution 40.9 the υ-value is 5~5.
He held it at 6, but occasionally shifted. To the clear solution, add NaC275,! Add ir, 4-(4'-[4-
1: 10-6-(4-to-1,1-methylammonio)anilino-s-triazin-2-ylaminoco-2'-methylphenylazo)penzamidinium dihydrochloride was filtered out and 50° It was dried to C.

Example 4 The method of Example 6 was applied to 4-aminophenyltrimethylammonium chloride (5.75%) and N-(2-7minoethyl)pyridinium chloride (3.2%) to F. ! It was repeated using The product obtained is 4-(4-(4-chloro-
6-(2-giridinoethylamino)-8-triazine-
2-ylaminocou 2'-methyl-phenylazo)penzamidinium dihydrochloride was obtained.

Example 5 A solution of 4-aminobenzamizone dihydrochloride 10.11 in 200 g of water and @ HCl 7.5.9 was cooled below 5°C and 2M Cydiazotization was carried out by adding 25 parts of NaN○2 solution. The solution was stirred for 10 minutes, the excess of nitrous acid was destroyed by adding a few drops of 10-thysulfamic acid, and
5.4 liters of N-methylaniline was added. - Add enough sodium acetate to raise the temperature to 5, and reduce the bath solution to 1
Stirred overnight. ffl, 4-(4-N-methylaminophenylazo)- added with NaCL60& stirred for 15 min.
The yellow precipitate of benzamidinium chloride was filtered off and
Dry at °C.

1 part of 4-(4-N-methylaminophenylazo)-benzamidinium chloride, 7.2 parts and 20 parts of methanol
0. V and water 200. Dissolved in a mixture of F. 4.71 of cyanuric chloride was added to the solution, which was cooled and kept below 5°C, and - if necessary, 25y of iM NaOH solution was added to maintain the solution at 5~B.4-(4-( The yellow precipitate of N-methyl-N-(4゜6-dichloro-8-triazin-2-yl)]aminophenylazo)benzamidinium chloride was filtered off, mixed with 1 g of mixed phosphate buffer and concentrated in vacuo. It dried inside.

The following table provides examples of other colored cationic compounds consistent with the first feature of the invention. These were obtained in the manner described in Example 4, when 75.4 g of N-methylamino were substituted for the amine equivalent weights given in Table 1.

Table 1 Example Amine 6 5-Methyl-2-methoxyaniline 7 6-acetoacetanili r 8 Acetoacet-4-aminoanilide 9 1-
4'-Aminophenyl-6-methyl-5-pyrazolone example 10 methanol 200. 4- in a mixture of F and 200 g of water.
6.6 g of (4'-N-methylaminophenyl a''j'')-benzamidinium chloride was dissolved. To the solution is added a solution of 2.1 g of 5-cyano-2,4,6-drichloropyridine in 10 g of acetone, - if necessary i M
The temperature was kept between -6 and 7 for 2 hours by adding 12 g of NaOH. At the end of this time, 4-(4-(:N-methyl-N-(5-cyano-4,6-cyclopyrimidin-2-yl)-aminocophenylazo)penzamidinium chloriP is filtered off. and dried in vacuo.

Example 11 11 g of 4-7 minofenylguanidinium dihydrochloride was added to a mixture of 200 g of water and concentrated HC17,5F. The solution was cooled to below 5°C and diazotized by adding 25.9 g of 2M NaN02 solution until an excess of nitrous acid was indicated by iodine starch paper. The solution was stirred for 10 minutes and the excess of nitrous acid was destroyed by adding a few drops of the acid solution to 10 thiosulfuric acid, followed by 5.4 drops of N-methylaniline.
g was added. Sodium acetate was added until -5, and the solution was stirred overnight, then NaCt 60. !
9 was added. After stirring for 15 minutes, 4-(4'-N-
The yellow precipitate of methylaminophenylazo)-phenylguanidinium chloride was filtered off and dried at 50°C.

4-( in a mixture of 200 g methanol and 200 g water
A solution of 7.2 g of 4'-N-methylaminophenylazo)-phenylguanidinium chloride was cooled to below 5°C, 4.7 g of cyanuric chloride was added, and - was added to I if necessary.
pH 5 for 2 hours by adding 425g of M NaC
It was maintained at ~6. 4-(4-CN-methyl-N-(4,
The yellow precipitate of 6-dichloro-8-triazin-2-yl)]aminophenylazo)-phenylguanidinium chloride was filtered off, mixed with 1 g of mixed phosphate buffer and dried in vacuo.

Example 12 4-(4-C2,4-dichloro-8-)riazine-6-
ylaminoco-2-methylphenyl)-azobenzamidinium chloride in chloroform-methanol-acetic acid (
The ff was prepared by chromatography on a silica column with a solution of i6:4:1 volume i), evaporating the solution and washing with methanol, acetone, aqueous acetone and finally water. The purified material is freeze-dried, which is 3
139001 at 90 nm. It had an absorption of not-'d1. 8g of a 9:1 mixture of dimethyl sulfoxide and water
4.5 g wet weight of agarose (CL-Sepharose 4B; pre-prepared with 500 g of water) in a solution of 0.1.
F, then a 1:1 mixture of dimethyl sulfoxide and water 1
00 g and finally washed with a 9:1 mixture of dimethylsulfoxy and water) were added followed by 0.02.9 g of solid lithium hydroxide and the mixture was rocked for 5 days. At the end of this time, the dyed del was filtered off and kept wet. Measurement of absorbance at c390 nm showed that the agarose contained 1.2 micromoles of bound dye per gram of wet del weight. An 8 mxx 20 mx column was packed with 1 g of the del and sodium acetate buffer (with a value of -4)
Buffer A) or -3-(N-morpholino) with value 7
Equilibration was achieved by washing with propane sulfonic acid buffer (buffer B). The buffers used are shown in Table 2.

containing appropriate units of enzymatic activity (trypsin, 1400: chymotrypsin, 4.24; carboxypezotidase B, 1.3), buffered with the equilibration buffer used previously.
6:) Thrombin, 24; Lactate dehydroke9nase,
1.2; and albumin, l], 42 A28o units)
A capacity of 20-150 Da was applied to the column. Unadsorbed proteins are washed from the column with equilibration buffer 5-7y, and then the adsorbed proteins are eluted with 0-7y over 10g of eluent.
This is done by separating I@ with a linear gradient solution of potassium chloride increasing to 1M. 1 g fractions were collected at a flow rate of 24 g/h and assayed for enzyme activity. The results are listed in Table 2.

Table 2 Albumin (human serum) Ba2O Lactate dehydrogenase A 65 0 (rabbit muscle) Chymotrypsin B 100 0
(Bovine pancreas) Thrombin (human plasma) B 0 87
0.3M KCl*In this example, zinc chloride 1
mM was added to the acetate buffer used.

Example 16 A suspension of 1 y of bovine pancreatic acetone powder in 10 g of 0.1 M acetate buffer, P) (4), was rocked overnight at 4°C and 25
After centrifugation at 00 rpm for 1 minute, the supernatant was filtered through a microporous cellulose prefilter. Trypsin 1rn9 was added to the supernatant. -0.5 M Tris/H
Ct buffer was adjusted to 7.9 and calcium chloride was added to a final concentration of 3 Q mM. After about 6 hours, a potency assay showed that maximum enzyme capacity was reached, at which point - was dialyzed. The dialyzed crude extract was filtered and now contains trypsin (activity: 5600 units) and chymotrypsin (activity: 1
A sample containing 40 units) was prepared as described in Example A and loaded with dye-agarose glue prepared as described in Example A and equilibrated in 0.1 PH5,5 acetate buffer.
Applied to a 1tm column. Unadsorbed proteins were washed from the column with acetate buffer 41.F, followed by 24.6 g of the same buffer containing 0.9M sodium chloride. Finally, the bound trypsin was eluted from the column with 5QnM glycine/Hct buffer 24.6.F, pH 2.1. Fractions weighing 8.2 g were collected at a flow rate of 9 B&/h and assayed for trypsin and chymotrypsin efficacy;
The ninth fraction collected had a total yield of 79 units and a specific activity of 7,400 units/m, which was 11 times stronger.

Claims (1)

[Claims] 1. A colored compound having a cationic group and a cellulose-reactive group. 2. General formula A-Y-B I or Y'-BII [wherein A represents a cationic group, Y represents a chromophore, Y' represents a cationic chromophore, and B represents a reactive group] The compound according to claim 1, which is a compound of 3. The reactive group is dichloropyrimidinyl, difluoropyrimidinyl, trichloropyrimidinyl, 5-chlorodifluoropyrimidinyl, 5-cyanodichloropyrimidinyl,
3. A compound according to claim 1 or 2, which is a mono- or poly-halogenoheterocycle selected from dichlorotriazinyl and monochlorotriazinyl. 4. The cationic group has the formula (i): a) ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼, b) ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ or c) ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ [Formula where R^1, R^2, R^3 and R^4 represent optionally substituted alkyl, alkenyl or aryl groups, B represents an aromatic system, D represents an alicyclic system, and At least one of R^1, R^2, R^3, R^4, B and D has a free value] and an acyclic or cyclic fully substituted ammonium group of formula (ii): ▲Math. , chemical formulas, tables, etc.▼ [In the formula, R^6, R^7, R^8 and R^9 represent H or an optionally substituted alkyl, alkenyl or aryl group, and n is 0 or 1. A compound according to any one of claims 1 to 3, which is selected from amidinium or guanidium groups. 5, R^1, R^2, R^3, R^4, R^6, R^7
, R^8, R^9, D and at least one of B is a chromophore. 6. The compound according to any one of claims 2 to 5, wherein the chromophore is an azo chromophore. 7. The cationic chromophore is 4-(3- and 4-trimethylammoniophenylazo)-aniline, 4-(3- and 4-guanidinophenylazo)-3-methylphenylamino, 4-(3'- and 4'-amidinophenylazo)phenylamino, 4-(3- and 4-amidinophenylazo)-3-methylphenylamino, 4-(3- and 4-amidinophenylazo)-2,5-
Dimethylphenylamino, 4-(3- and 4-amidinophenylazo)-5-methyl 1-2-methoxyphenylamino, 4-(3- and 4-amidinophenylazo)-2,5-
Dimethoxyphenylamino, 4-(3- and 4-amidinophenylazo)-3-acetylaminoaniline, 4-(3- and 4-amidinophenylazo)-3-ureidophenylamino, 4-(3- and 4- amidinophenylazo)-6-methoxy-3-acetylaminophenylamino, 4-(3- and 4-amidinophenylazo)-N-methylphenylamino, 4-([4-guanidinophenylazo]-[acetyl]
-acetylamino)-phenylamino, 4-[4-(4-trimethylammoniophenylazo)
-3-methylpyrazol-5-one-1-yl]-phenylamino, 4-[4-(4-amidinophenylazo)-3-methylpyrazol-5-one-1-yl]-phenylamino,
and 4-[4-(4-guanidinophenylazo)-3-methylpyrazol-5-one-1-yl]-phenylamino.