Photochromism can be defined as the property of a material able to change reversibly its visible absorption spectrum upon exposure to activating radiation and to revert to its original absorption spectrum thermally on removal of the activating radiation, or on exposure to radiation of a different wavelength.

US Patent No. 4220708 describes a series of photochromic succinic anhydride and succinimide derivatives (fulgides and fulgimides respectively) having the following general formula (1): (I) wherein X represents 0, NR6, R6 being H, alkyl, aryl or an aralkyl group; R represents an alkyl, aryl, aralkyl, or heterocyciic group; A represents a 3-furyl, 3-thienyl, 3-benzofuryl or 3-benzothienyl group; B represents a cycloalkylidene group or the group,

in which R2 and R3 independently represent an alkyl, aryl, aralkyl or a heterocyclic group, or one of R2 and R3 represents hydrogen and the other represents an alkyl, aryl, aralkyl, or heterocyclic group.

Compound of this class undergo cyclisation when exposed to ultraviolet radiation, ring closure taking place between the carbon atom to which groups R2 and R3 are attache and the 2-position of the furyl or thienyl ring. The photocyclisation rection for fulgide ring structures is illustrated by the following scheme (Scheme 1).

Scheme 1 The cyclic forms of the fulgides, for example (III), are highly coloured, usually in the bright red to deep purple range and this is reporte to arise from the extended, near plana, conjugated double bond structure with the oxygen heteroatom at one end of the molecule and the conjugated carbonyl at the other.

Compound of formula (I) in which X is oxygen may be converted into the corresponding fulgimide derivatives (1, X = NR6) by rection of the anhydride with a primary amine followed by an acid chloride in a suitable solvent such as dichloromethane.

As with the fulgides, fulgimides undergo photochemical ring closure to form the more highly coloured, thermally stable product. They exhibit photochromic properties similar to the corresponding fulgides, except that the long wavelength (visible) absorption bands of the coloured forms are generally broader and show bathochromic shifts. Furthermore fulgimides display greater resistance to hydrolysis compare with fulgides. Suitable fulgimide derivatives may be coupled with target biological systems, thereby giving such a system photochromic properties. For example, a fulgimide derivative in which R CH2COOH was coupled via its N-hydroxysuccinimide ester to the nucleotide analogue, aminoallyl-2'-deoxyuridine 5'-triphosphate (V. Kiruvanayagam, PhD Thesis, University of Wales, (1995), p137).

Replacement of one or the other of the carbonyl groups of the fulgide structure with a substituted nitrogen results in two alternative isomers of fulgimides, termed a-isofulgimides (IV) and ß-isofulgimides (V). The compound undergo photochemical ring closure to give coloured forms.

R=CH2Ph<BR> <BR> <BR> <BR> <BR> α-Isofulgimideß-Isofulgimide (IV) (V) a-lsofulgimides may be prepared by the rection of the appropriate succinic half-ester derivative with the Grignard derivative of the required amine, followed by cyclisation with DCC. ß-lsofulgimides can be prepared by a similar method,

but a more convenient approach is to derive the required succinamic acid from the corresponding fulgide, followed by rection with DCC (K. S. V. Koh, PhD Thesis, University of Wales, 1993).

Accordingly the present invention provides compound of formula VI: or stereoisomers thereof, wherein groups B'and B2 are selected from the groups: such that B'w B2; R7, R8 and R9 are hydrogen or are chosen to provide desired solubility, reactivity and spectral properties to the compound; X is a group- (CH2) n- or a group: in which case it links via a two or three atom chain with carbon atom Ca of ring Z to form a fused bicyclic aromatic ring which may be optionaíly substituted and n is O or 1;

Z is an optionally substituted six-membered aromatic or fused bicyclic aromatic ring containing carbon atoms and optionally no more than two atoms selected from oxygen, nitrogen and sulphur; R'and RZ independently represent an alkyl, cycioalkyl, aryl, or an aralkyl group or one of R'and R2 represents hydrogen and the other an alkyl, cycloalkyl, aryl, or an aralkyl group, or the group: represents an adamantylidene group; R3 represents hydrogen, alkyl, or aryl; A represents a substituted or unsubstituted heterocyclic ring having one of the following structures: where R4 is selected from hydrogen, alkyl, aryl, and aralkyl groups, and R5 is selected from hydrogen, C1-12 hydrocarbyl optionally substituted with halogen, Cl-6 alkoxy, aryt and Cs-, 2 aryloxy groups and E is selected from 0, S and NR6, where R6 is selected from hydrogen, C 1-6 alkyl, aryl or an aralkyl group.

Suitably, the two or three atom chain which links group X with carbon atom Ca of ring Z contains carbon atoms and optionally no more than one nitrogen atom.

Preferably the two or three atom chain contains carbon atoms.

Optional substituents R7, R8 and R9 on the aromatic ring Z or the bicyclic aromatic ring that incorporates Z are the same or different and are independently selected from-R'° and-L-R'°, wherein R10 is selected from:

neutral groups that reduce water solubility; polar groups that increase water solubility; target bonding groups such as functional groups that can be used in labelling rections; reactive groups; electron donating and withdrawing groups that shift the emission wavelengths of the photochromic molecule; lipid and hydrocarbon solubilising groups, and L is selected from the group consisting of a straight or branche C1-10 alkyl chain, a C2-: o monoether or polyether and a C2-10 atom chain containing up to two secondary amide linkages.

Preferred R'° groups are selected from: hydrogen, halogen, amide, Cl-C6 alkoxy, cyano, aryl, heteroaryl, sulphonate, quaternary ammonium, hydroxyl, optionally substituted amino, sulphydryl, carbonyl, and reactive groups, for example, succinimidyl ester and anhydride, and groups reactive with amino, hydroxyl, carboxyl, aldehyde, or sulphydryl groups.

Specific examples of the reactive groups R7, R8 and R9 and the groups with which R7, R8 and R9 will react are provided in Table 1. In the alternative, the R7, R3 and R9 may be the functional groups of Table 1 which would react with the reactive groups of a target molecule.

Table 1: Possible Reactive Substituents and Functional Groups Reactive Therewith Reactive Groups Functional Groups succinimidyl esters primary amino, secondary amino anhydrides primary amino, secondary amino, hydroxyl substituted hydrazines, aldehydes, ketones acid halides amino groups haloacetamides, maleimides thiols, imidazoles, hydroxyl, amine carbodiimides carboxyl groups phosphoramidites hydroxyl groups

Preferred reactive groups R7, R8 and R9 which are especially useful for labelling target components with available amino and hydroxyl functional groups include: where n is 0 or an integer from 1-10.

In one preferred embodiment of the present invention the compound of formula (VI) have the formula (Vla): or a stereoisomer thereof, wherein groups R', R2, R3, R7, R8, R9, A, X and Z are hereinbefore defined.

In a second preferred embodiment of the present invention the compound of formula (VI) have the formula (Vlb):

or a stereoisomer thereof, wherein groups R', R2, R3, R', R8, R9, A, X and Z are hereinbefore defined.

In one embodiment of the present invention R6 is selected from hydrogen, C 1-6 alkyl, or aralkyl group and L is selected from the group consisting of a straight or branche Cl-io alkyl chain, and a C2-10 monoether or polyether.

Alkyl is a straight or branche chain alkyl group containing from 1-20 carbon atoms, preferably 1 to 6 carbon atoms, for example methyl, ethyl, n-propyl, iso- propyl and butyl.

Aryl is an aromatic substituent containing one or two fused aromatic rings containing 6 to 10 carbon atoms, for example phenyl or naphthyl, the aryl being optionally and independently substituted by one or more substituents, for example halogen, straight or branche chain alkyl groups containing 1 to 10 carbon atoms, cycloalkyl, aralkyl and alkoxy for example methoxy, ethoxy, propoxy and n-butoxy.

Cycloalkyl is an alicyclic substituent containing from 3 to 6 carbon atoms being attache by a single bond, for example cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

Heteroaryl is a mono-or bicyclic 5 to10 membered aromatic ring system containing at least one and no more than 3 heteroatoms which may be selected from N, 0, and S and is optionally and independently substituted by one or

more substituents, for example halogen, straight or branche chain alkyl groups containing 1 to 20 carbon atoms, cycloalkyl, aralkyl and alkoxy for example methoxy, ethoxy, propoxy and n-butoxy.

Aralkyl is a Cl to C6 alkyl group substituted by an aryl or heteroaryl group.

Halogen and halo groups are selected from chlorine, bromine and iodine.

For the purpose of increasing water solubility or reducing unwanted non- specific binding of the photochromic compound of the present invention to inappropriate components of a sample, one or more of the R7, R3 and R9 groups may be selected from well known polar or electrically charged chemical groups.

Examples of such groups are-S-F, where F is hydroxy, sulphonate, carboxylate, substituted amino or quaternary amino and where S is a spacer group such as -(CH2) n-where(CH2) n-where n is 0 to 6. Examples of-S-F groups include C1-6 alkyl sulphonates, such as- (CH2) a-SOs and- (CH24-SO3.

Specific examples of the compound of the pèsent invention are as follows: iy E-4-Dicyclopropylmethylene-3-f1- (2, 5-dimethyl-3-furyllethylidene- benzimidazol [1, 2-alpyrrolidin-2-one ; ii) E-4-Diphenylmethylene-3- (1- (2, 5-dimethyl-3-thienyl) ethylidenel- benzimidazol (1, 2-al pyrrolid i n-2-one; iii)E-4-Dicyclopropylmethylene-7,8-dimethyl-3-[1-(2-methyl-5 -phenyl-3- thienyl)ethylidene]benzimidazol[1,2-a]pyrrolidin-2-one; iv) E-3-Adamantylidene-7,8-dimethyl-4-[1-(2,5-dimethyl-3-furyl)e thylidene]- benzimidazol (1, 2-aJpyrrolidin-2-one; v) E-3-Adamantylidene-4-[1-(2,5-dimethyl-3-furyl)ethylidene]- benzimidazolf1, 2-alpyrolidin-2-one-8-carboxylic acid; vi) E-8-Amino-4-dicyclopropylmethylene-3- [1- (2, 5-dimethyl-3- furyl)ethylidene]benzimidazol[1,2-a]pyrrolidin-2-one;

vii) Z-4-Adamantylidene-8-nitro-3-f1- (2, 5-dimethyl-3- furyl)ethylidenelbenzimidazol f 1, 2-alpyrrolidin-2-one; viii) Phenacyl Z-3-dicyclopropylmethylene-4-t1- (2,5-dimethyl-3- furyl) ethylidenelbenzimidazolf 1, 2-alpyrrolidin-2-one-8-carboxylate; ix) E-3-Dicyclopropylmethylene-4- (1- (2,5-dimethyl-3- furyl) ethylidenelbenzimidazolf 1, 2-alpyrrolidin-2-one-8-carboxylic acid.

The groups provided herein are not intended to be all-inclusive of those groups which can be incorporated at the R7, R8 and R9 sites of the present invention. It will be understood that there are various other groups which will react with groups on material that is to be labelle by the compound of the present invention. Compound produced by the incorporation of other such groups at the R7, R8 and R9 sites are intended to be encompassed by the present invention.

The compound of the present invention may be used in one or more biological and non-biological applications. With respect to non-biological applications, compound of the present invention having one or more uncharged groups at the R7, R8 and R9 positions, for example alkyl and aryl moities, may be dissolve in non-polar materials to provide photochromic properties to those materials. Such non-polar materials inclue, for example paints, polymers, plastics, waxes, oils, inks and hydrocarbon solvents. Another non-biological application of the photochromic compound of the present invention is to dissolve compound having one or more charged and/or polar groups at the R, R8 and R9 positions in polar solvents such as water, alcools such as methanol or ethanol, ethylene glycol, or mixtures of such solvents.

Alternativeíy, the photochromic compound of the present invention may contain a polymerizable group suitable for the formation of a polymer containing the complex. Suitable polymerizable groups are selected from acrylate, methacrylate and acrylamide. Polymerization may be carried out with a

suitably derivatized compound of this invention used in conjunction with a second polymerizable monomer starting material, such as styrene or vinyltoluene, to form a copolymer containing the photochromic compound. The photochromic compounds need not have a polymerisable group, for example, the complex may be incorporated during polymerisation or particle formation or may be absorbe into or onto polymer particles.

Compound of the present invention having a functional or reactive group at the R7, R8 and R9 positions may be used to covalently label a target mateFial to impart photochromic properties to the target material, such as a carrier material, a luminescent compound or a biological material. The target bonding group may be a reactive group for reacting with a functional group on the target material. Alternatively the target bonding group may be a functional group for rection with a reactive group on the target.

Suitable target materials may include a luminescent compound, such as fluorescent dyes (based for example on the fluorescein, rhodamine, coumarin, pyrene and cyanine chromophores), antibodies, antigens, proteins, carbohydrates, lipids, nucleotides which contain or are derivatized to contain one of amino, hydroxyl, sulphydryl, carboxyl, or carbonyl groups, and oxy or deoxy polynucleic acids which contain or are derivatized to contain one of amino, hydroxyl, phosphate, thiophosphoryl, sulphydryl, carboxyl, or carbonyl groups, cells, polymer particles, or glass beads. Covalent labelling using compound of the present invention may be accomplished with a target having at least one functional or reactive group as defined hereinbefore. The target may be reacted with an amount of a compound of the present invention having at least one of R7to R9 that inclues a reactive or functional group as hereinbefore defined that can covalently bind with the functional or reactive group of the target material. The target material and the compound of the pèsent invention are incubated under conditions and for a period of time

sufficient to permit the target materai to covalently bond to the compound of the present invention.

The present invention also provides a process for the preparation of a compound of formula (Vl) which comprises rection of a compound of formula (VII), or its corresponding di-carboxylic acid, di-Ca-C6 alkyl ester, or mono-carboxylic acid-mono Cl-C6 alkyl ester derivative, wherein R, R2, R3 and A are as hereinbefore defined with a compound of formula (vil): or a salt thereof, optionally substituted by groups R7, R8 and R9, wherein R', R8 and R9, X and Z are as hereinbefore defined. The rection is suitably carried out in a dry, inert solvent such as toluene and in the absence of light. The rection is suitably carried out at an elevated temperature, for example 50°C to 150°C, suitably 100°C to 125°C. The rection mixture (containing a mixture of

geometric isomers of the product) is fractionated into separate isomers by column chromatography using silica gel, or by fractional crystallisation.

The Stobbe condensation provides a general method for preparing compound of the general formula (VII). An account of this rection and its application to the synthesis of a wide range of succinic acid derivatives is given in Organic Rections, Vol. 6, pp1-73, published by Wiley, New York 1951. For example, compound of formula (VII) can be prepared by rection of a compound of formula (IX), wherein R3 and A are as hereinbefore defined with a succinic ester of formula (X) I where R'and R 2 are as hereinbefore defined and R'and R"are independently selected from methyl, ethyl, n-propyl, and n-butyl, by a Stobbe condensation to yield a product of formula (VII). Preferably the Stobbe condensation may be carried out by treating the reactants in t-butanol, or toluene, or tetrahydrofuran containing potassium t-butoxide. The product at this stage is the half-ester, ie where one of the R'or R"groups is hydrogen. The half-ester is then converted into the di-acid by hydrolysis, for example by boiling with ethanolic potassium

hydroxide. The di-acid is then converted into its anhydride by a dehydration rection, for example by stirring at ambient temperature with an acid chloride.

Preferably acetyl chloride is used for this purpose.

For example: (i) Compound of formula (VII) can be prepared using as the starting material a ketone of formula (XI), wherein R3, R4, Rs and E are as hereinbefore defined with a succinic ester of formula (X), where R', R2, R'and R"are hereinbefore defined.

(ii) Compound of formula (VII) above can also be prepared in an analogous manner to that described above using adamantan-2-one as a starting material.

Preparation of adamantan-2-one is described in US Patent No. 3257456 and by Gulak et al, Organic Synthesis, 53,8, (1973). Compound of formula (VII) can be prepared by refluxing adamantan-2-one with a succinate diester in a solution of potassium t-butoxide in t-butanol to give the potassium salt of the corresponding half ester, which is converted into an adamant-2-ylidene succinate diester of formula (XII), in which R'and R"are hereinbefore defined.

Compound of formula (VII) containing an adamantylidene group are obtained by reacting a diester of formula (Xll) with a ketone of formula (IX) or of formula (XI) in the presence of sodium hydrie or potassium t-butoxide, in a dry inert solvent such as toluene. See for example, US Patent No. 4220708, the disclosure of which is incorporated by reference.

Precursor compound of formula (vil) are readily available or may be prepared by methods well known to those skilled in the art.

It will be readily appreciated that certain compound of formula (Vl) may be useful as intermediates for conversion to other compound of the formula (VI) by methods well known to those skilled in the art. Likewise, certain of the intermediates may be useful for the synthesis of derivatives of formula (VI).

The compound of the present invention may be synthesized by the methods disclosed herein. Derivatives of the compound having a particular utility are prepared either by selecting appropriate precursors or by modifying the resultant compound by known methods to include functional groups at a variety of positions. As examples, the compound of the present invention may be modifie to include certain reactive groups for preparing a photochromic labelling ragent, or charged or polar groups may be added to enhance the solubility of the compound in polar or nonpolar solvents or materials. As examples of conversions, carboxylic acid groups may be converted into esters, and amide groups.

Compound of the present invention switch from near colourless to coloured forms on exposure to UV light and the colour of the coloured form is pH dependent.

The invention is further exemplified by reference to the following examples and Figure 1 which illustrates a fluorescence study of a photochromic compound- fluor switch according to Example 11.

Experimental Section i) General Ultraviolet spectra were recorde on a Cecil CE6600 spectrophotometer and a Perkin Elmer Lambda 20 UV/Vis spectrophotometer, using UV Winlab, for 1 x 10-4M solutions in dry toluene.

'H NMR spectra were obtained using a Bruker WM 400 (400MHz) or a Bruker WM 360 (360MHz) FT NMR spectrometer for samples in deuterated chloroform with 1 % TMS as internal standard.

Microanalyses were obtained using a Perkin Elmer 240B analyser.

Melting points were measured on a Reichert Hot Stage Microscope.

Photochemical rections employed an optical bench incorporating a focussed medium pressure 100W mercury arc lamp and either a Woods glass OXO A filter to produce 366nm light or a 370nm cut-off yellow filter for bleaching.

ii) General Synthesis of Compound of Formula (VI) A solution of a compound of formula (VII) and a compound of formula (vil) in toluene was boiled (24 houris) in the absence of light. Solvent was removed under reduced pressure and the residual oil was purifie by column chromatography on silica gel, using usually mixtures of diethyl ether and petroleum ether (40-60°C) as eluant. The main fractions gave compound of formula (VI) usually as yellow crystals after recrystallisation from diethyl ether and petroleum ether (40-60°C).

Example 1: Preparation of Compound 1 a, 1 b and 1 c A solution of E/Z-3-dicyclopropylmethylene-4-f 1- (2, 5-dimethyl-3- furyl) ethylidenelsuccinic anhydride (5. OOg, 16. 03mmol) and 1,2- phenylenediamine (2.08g, 19.23mmol) in toluene (50ml) was boiled for 23 hours. Removal of solvent left an oil consisting of a mixture of compound of formula 1 a, 1 b and 1 c, which were separated and purifie by column chromatography and by fractional recrystallisation from diethyl ether and (40-60°C).petroleumether

1a:E-4-Dicyclopropylmethylene-3-[1-(2,5-dimethyl-3-furyl)eth ylidene]- benzimidazolf1, 2-alpyrrolidin-2-one Yellow crystals (0.81g, 13%), m. p. 148-149°C, m/z 384.2. Found: C, 78.15; H, 6.22; N, 7.19%. C26H24N202 requires C, 78.10; H, 6.25; N, 7.29%.

1 b: Z-3-Dicyclopropylmethylene-4-[1-(2,5-dimethyl-3-furyl)ethyli dene]- benzimidazol[1,2-a]pyrrolidin-2-one Yellow crystals (0. 65g, 11%), m. p. 192-194°C, m/z 384.2. Found: C, 77.92; H, 6.44 ; N, 7.13%. C2rH24N202 requires C, 78.10; H, 6.25; N, 7.29%.

1c:Z-4-Dicyclopropylmethylene-3-[1-(2,5-dimethyl-3-furyl) ethylidene]- benzimidazol[1,2-a]pyrrolidin-2-one Yellow crystals (0.25g, 4%), m. p. 214-216°C, m/z 384.2. Found: C, 78.05; H, 6.37; N, 7.04%. C26H24N202 requires C, 78.10; H, 6.25; N, 7.29%.

Example 2: Preparation of Compound 2 A solution of E-[1-(2, 5-dimethyl-3-thienyl) ethylideneldiphenylmethylene succinic anhydride f0. 50g, 1.25mmol) and 1,2-phenylenediamine (0.16g, 1.48mmol) in toluene (30ml) was boiled for 68 hours. Removal of solvent left an oil

containing the impure compound of formula 2, which was purifie by column chromatography and by fractional recrystallisation from diethyl ether and petroleum ether (40-60°C).

2: E-4-Diphenylmethylene-3- [l- (2,5-dimethyl-3-thienyl) ethylidenel- benzimidazol(1, 2-al pyrrolidin-2-one Yellow crystals (0.25g, 42% 1, m. p. 224-225.5°C, m/z 472.1. Found: C, 78.78; H, 5.32; N, 6. 01%. C31H24N2OS requires C, 78.79; H, 5.12; N, 5@93%.

Example 3: Preparation of Compound 3a and 3b A solution of E-3-dicyclopropyimethylene-4- (1- (2-methyl-5-phenyl-3- thienyl)ethylidene] succinic anhydride (1. OOg, 2.56mmol) and 4,5-dimethyl-1,2- phenylenediamine f0. 42g, 3. 08mmol) in toluene (40ml) was boiled for 69 hours. Removal of solvent left an oil consisting of a mixture of compound of formula 3a and 3b, which were separated and purifie by column chromatography and by fractional recrystallisation in diethyl ether and petroleum ether (40-60°C).

3a: E-4-Dicyclopropylmethylene-7,8-dimethyl-3-[1-(2-methyl-5-phe nyl-3- thienyl)ethylideneJbenzimidazolf 1, 2-alpyrrolidin-2-one Green/yellow crystals (0. 16g, 13%), m. p. 213. 5-215.5°C, m/z 490.0. Found: C, 78.30; H, 6.20; N, 5.65%. C32H3oN20S requires C, 78.33; H, 6.16; N, 5.7 1 %.

3b: Z-3-Dicyclopropylmethylene-7, 8-dimethyl-3-fl- (2-methyl-5-phenyl-3- thienyl)ethylidene]benzimidazol[1,2-a]pyrrolidin-2-one Pale yellow crystals (0.28g, 22%), m. p. 233-236°C, m/z 490. 1. Found: C, 78.12; H, 6.34; N, 5.78%. C32H3oN20S requires C, 78.33; H, 6.16; N, 5.71 %.

Example 4: Preparation of-Compounds 4a and 4b A solution of E-3-adamantylidene-4-f 1- (2, 5-dimethyi-3-furyliethylidenelsuccinic anhydride (1. OOg, 2.84mmol) and 4,5-dimethyl-1,2-phenylenediamine l0. 58g, 4.26mmol) in toluene (40ml) was boiled for 22 hours. Removal of solvent left an oil consisting of a mixture of compound of formula 4a and 4b, which were separated and purifie by column chromatography and by fractional recrystallisation in diethyl ether and petroleum ether (40-60°C).

4a: E-3-Adamantylidene-7,8-dimethyl-4-[1-(2,5-dimethyl-3-furyl)e thylidene]- benzimidazolf1, 2-alpyrolidin-2-one Yellow/Green cube shaped crystals (0.26g, 20% 1, m. p. 243-244°C, m/z 452.3.

Found: C, 79.84; H, 6.91; N, 6.09%. C3oH32N202 requires C, 79.61; H, 7.13; N, 6.19%.

4b: E-4-Adamantylidene-7,8-dimethyl-3-[1-(2,5-dimethyl-3-furyl)e thylidene]- benzimidazol[1,2-a]pyrrolidin-2-one Colourless crystals (which turned blue on surface) (0. 089, 6%), m. p. 221- 223°C, m/z 452.3. Found: C, 79.35; H, 7.25; N, 6.24%. C3oH32N202 requires C, 79.61; H, 7.13; N, 6.19%.

Example 5: Preparation of Compound 5 A solution E-3-adamantylidene-4- [1- (2, 5-dimethyl-3-furyl) ethylidenelsuccinic anhydride (1. OOg, 2.84mmol), 3,4-diaminobenzoic acid (0.52g, 3. 42mmol) and a catalytic amount of potassium tert butoxide (0.032g, 2. 85#10-4mol) in toluene (30ml) were boiled for 49 hours. Removal of solvent left an oil containing the impure compound of formula 5, which was purifie by column chromatography using ethyl acetate and petroleum ether (40-60°C) followed by fractional recrystallisation in a hot 50%: 50% toluene/ethanol solution.

5: E-3-Adamantylidene-4-[1-(2,5-dimethyl-3-furyl)ethylidene]ben zimidazol- t1, 2-alpyrrolidin-2-one-8-carboxylic acid Yellow/white powder (0. 16g, 12%), m. p. 271-273°C, m/z 468.2.

Found: C, 74.12 ; H, 6.15; N, 5.68%. C29H28N2O4 requires C, 74.34; H, 6.02; N, 5.98%.

Example 6: Preparation of Compound 6a and 6b A solution of E/Z-3-dicyclopropylmethylene-4- [l- (2,5-dimethyl-3- furyl) ethylidenelsuccinic anhydride (1.50g, 4. 81mmol) and 4-amino-1,2- phenylenediamine, fade by the catalytic hydrogénation of 4-nitro-1,2- phenylenediamine (2. OOg, 13.07mmol) using Raney nickel and hydrazine (2ml), in toluene (40m1) 1 was boiled for 96 hours. Removal of solvent left an oil consisting of compound of formula 6a and 6b, which were separated and purifie by column chromatography and by fractional recrystalíisation in diethyl ether and petroleum ether (40-60°C). Compound 6b was obtained admixed with small amounts of isomers.

6a: E-8-Amino-4-dicyclopropylmethylene-3-[1-(2,5-dimethyl-3- fuyl) ethylidenelbenzimidazolf 1, 2-alpyrolidin-2-one Yellow/brown powder (0.11 g, 6%), m. p. 91-94°C, m/z 399.4. Found: C, 74.96; H, 6.54; N, 10.71%. C25H25N3O2 requires C, 75.19; H, 6.27; N, 10.53%.

6b Z-8-Amino-3-dicyclopropylmethylene-4-[1-(2,5-dimethyl-3- furyl) ethylidenelbenzimidazol [1,2-alpyrrolidin-2-one Green powder (0.13g, 7%), m/z 399.5. Found: C, 74.92; H, 6.52; N, 10.62%.

C2sH2sN302 requires C, 75. 19; H, 6.27; N, 10.53%.

Example 7: Preparation of Compound 7 A solution of methyl Z-2-adamantylidene-3-[1-(2, 5-dimethyl-3-furyl) ethylidenel succinate (0.75g, 1.95mmol) containing 4-nitro-1,2-phenylenediamine (0.33g, 2.15mmol) was boiled in dichloromethane (40ml) with 1-ethyl-3- (3- dimethylamino) propylcarbodiimide hydrochloride (0.82g, 4.28mmol) for 18 hours. Removal of solvent left an oil containing the impure compound of formula 7, which was purifie by column chromatography using diethyl ether and petroleum ether (40-60 °C), followed by fractional recrystallisation from dichloromethane and petroleum ether (40-60 °C).

7: Z-4-Adamantylidene-8-nitro-3-[1-(2,5-dimethyl-3-furyl)ethyli dene] benzimidazolf 1, 2-alpyrrolidin-2-one Yellow needles (0.20g, 22%), m. p. 232-234 °C, m/z 470.4.

Example 8: Preparation of Compound 8 A solution of Z/E-4-methyl 2-dicyclopropylmethylene-3- (1- (2, 5-dimethyl-3-furyl) ethylidenelsuccinates (4.50g, 13.08mmol) containing phenacyl 3,4-diamino- benzoate (3.50g, 12. 96mmol) was boiled in dichloromethane (50ml) with 1- ethyl-3-(3-dimethylamino) propylcarbodiimide hydrochloride (5.50g, 28.72mmol) for 3 hours. Removal of solvent left an oil containing impure compound of formula 8, which was purifie by column chromatography using ethyl acetate and petroleum ether (40-60 °C), followed by fractional recrystallisation from dichloromethane and (40-60°C).ether 8: Phenacyl Z-3-dicyclopropylmethylene-4- (1- (2, 5-dimethyl-3- furyl)ethylidene]benzimidazol[1,2-a]pyrrolidin-2-one-8-carbo xylate Yellow crystals (1.34g, 18%), m. p. 213-215 °C, m/z 547.1.

Example 9: Preparation of Compound 9 Phenacyl Z-3-dicyclopropylmethylene-4- [1- (2, 5-dimethyl-3-furyl) ethylidenel benzimidazolf1, 2-a) pyrrolidin-2-one-8-carboxylate (2.50g, 4.58mmol) was dissolve in dry dichloromethane (20ml) containing glacial acetic acid (1 Oml) and stirred. Zinc dust (9g, 0.14mol) was added over a period of 10 minutes and then stirred for 30 minutes. The rection mixture was filtered, and the filtrate washed with water (4 x 200ml) and dried over anhydrous sodium sulphate. Removal of solvent left an oil containing impure compound of formula 9, which was purifie by column chromatography using ethyl acetate and petroleum ether (40-60 °C), followed by fractional recrystallisation from diethyl ether and petroleum ether (40-60 °C).

9: E-3-Dicyclopropylmethylene-4-[1-(2,5-dimethyl-3-furyl)ethyli dene] benzimidazolfl, 2-alpyrrolidin-2-one-8-carboxylic acid Yellow/green powder (0.27g, 14%), rn. p. 234.5-235.5 °C, m/z 428.3. Found: C, 72.74; H, 5.89; N, 6. 27%. C26H24N2O4 requires C, 72.88; H, 5.65; N, 6.54%.

Example 10: Preparation of Compound 10 A solution of E-3-dicyciopropylmethylene-4- (1- (2, 5-dimethyl-3-furyl) ethylidenel benzimidazol[1,2-a]pyrrolidin-2-one-8-carboxylic acid (0.30g, 7.01 X 10-4MOI) in dichloromethane (20m1 ! was stirred under nitrogen with thionyl chloride (0. 20ml, 0.33g, 2.74mmol) for 10 minutes. Solvent and excess thionyl chloride was removed on a rotary evaporator. Under nitrogen atmosphere and in the dark, the residual oil dissolve in dichloromethane (20ml) was added dropwise over a period of 1 hour, using a pressure equalising dropping funnel, to a stirred solution of ethylenediamine (0. 45ml, 0.40g, 6.73mmol) in dichloromethane (20ml). The rection mixture was washed with water (2 x 100ml) and dried. Petroleum ether (40-60 °C) was added and the solvent removed on a rotary evaporator until compound of the formula 10 separated out as a light brown powder (0.12g, 36%), m. p. 168-179 °C, m/z 471.4.

Example 11: Preparation of Photochrome-Cy3 conjugate (Compound 11)

To a stirred solution of bis-functionai suíphonated Cy3 dye (Amersham Pharmacia Biotech) (100mg, 1.49 x 10-4 mol) in acetonitrile (20ml) was added 1, 3-dicyclohexylcarbodiimide (59mg, 2.86 x 10~4mol) and N-hydroxysuccinimide (35mg, 3.04 x 10~4mol). The rection mixture was stirred in the dark for 18 hours under nitrogen. The rection mixture was filtered and solvent removed from the filtrate on a rotary evaporator leaving a purple solid (0.107g). A sample of this solid (53mg) was dissolve in dimethylformamide (7ml) and stirred in the dark under nitrogen. To this was added dropwise over 3 heurs, a solution of compound of formula 10 (28mg, 5.96 x 10-5mol) in dimethylformamide (2moi). The rection mixture was stirred for 17 hours and filtered. Solvent was removed from the filtrate on a rotary evaporator and the residual oil dissolve in a minimum amount of methanol and loaded onto a glass backed reverse phase (RP-18-254s) pre-coated preparative plate and separated using a 90% methanol: 10% water eluant system. The appropriate band (RF range 40%-50%) was scratche off the plate washed with methanol to extract product and filtered. Removal of solvents from the filtrate on a rotary evaporator gave compound of formula 11 as a purple solid (23mg).

Fluorescence study of photochromic compound-fluor switch (Scheme 2 and Figure 1) A solution of compound of formula 11 in methanol had its emission spectrum (Excitation 550 nm) measured before exposure to UV light (Figure 1, spectrum 1).

The emission spectrum (Excitation 550 nm) of the same sample was measured following irradiation (366 nm) for 2 minutes using a 12W hand lamp incorporating Woods glass filter (Figure 1, spectrum 2). (Note: exposure to UV light causes the ring closure of compound of formula 11 to compound of formula 12, (Scheme 2)).

The emission spectrum (#Exeitation 550 nm) of the same sample was measured following bleaching using five flashes from a flash gun mounted with a 370nm cut-off yellow filter (Figure1, spectrum 3). (Note: bleaching with visible light causes ring opening of compound of formula 12 to regenerate the compound of formula 11, (Scheme 2)). 03S S03 Spectrum I H ce+ /cl2+ \ I N od /O H O O 11 uv 03S S03 Spectrum 2 < N N N / 0 0 X N ft O 12 Visible 03S S03 Spectrum 3 ß s +I I N N/Caz+ \ \ I N 0 H /O H O 0 V 11