The use of dyes in modern biomedicine

The discovery of the aniline dyes in the 19th century and contemporary investigation of their use as biological stains by scientists such as Koch and Ehrlich led to the idea of selectivity and formed the basis of modern chemotherapy; several of these dyes remain in pharmacopoeias. While the development of therapeutics has tended to avoid colored compounds due to unwanted coloration, the modern application of photosensitizing dyes, both in the fields of cancer therapy and anti-infection, depends on this phenomenon. In addition, the fluorescence of some anticancer photosensitizers allows their use as tumor localizing agents, which is particularly useful in precancerous conditions. It is also fitting that dyes employed in Ehrlich's original studies, such as the phenothiazinium dye, methylene blue, are now in clinical use for disinfecting donated blood products.     

The use of dyes in modern biomedicine

The discovery of the aniline dyes in the 19th century and contemporary investigation of their use as biological stains by scientists such as Koch and Ehrlich led to the idea of selectivity and formed the basis of modern chemotherapy; several of these dyes remain in pharmacopoeias. While the development of therapeutics has tended to avoid colored compounds due to unwanted coloration, the modern application of photosensitizing dyes, both in the fields of cancer therapy and anti-infection, depends on this phenomenon. In addition, the fluorescence of some anticancer photosensitizers allows their use as tumor localizing agents, which is particularly useful in precancerous conditions. It is also fitting that dyes employed in Ehrlich's original studies, such as the phenothiazinium dye, methylene blue, are now in clinical use for disinfecting donated blood products.     

Microalgae as source of biofuel,food, fodder,and medicines

Current status and future prospects of such problem as the production of microalgae and their application for biofuel generation (biodiesel, biohydrogen, bioethanol), as well as other products, is discussed in the review. The use of microalgae in human food, fodder, cosmetics, dyes, polysaccharides, antioxidants, medicines, and other products is quite promising. Presently, microalgae are noncompetitive with plant materials, due to economic reasons, in serving as a source of biofuel. Thereby, it is urgently necessary in modern biotechnology to improve the methods for the production of microalgae and search for new ways of their processing.     

Comparison of historic Grübler dyes with modern counterparts using thin layer chromatography

The aniline dye industry was created in 1856 when William Perkin prepared the dye, mauve, from coal tar. Following that discovery, several dye manufacturing businesses were formed in Western Europe, most successfully in Germany. It was to these companies that early investigators turned to obtain these new dyes for the developing field of biology. In 1880, Dr. Georg Grübler started a company in Germany to supply the needs of biologists. Grübler dyes developed a reputation for excellence. In the study reported here, 29 samples of 12 Grübler dyes were compared to modern counterparts using thin layer chromatography. The dyes studied were basic fuchsine, acid fuchsine, safranine, pyronine, aniline blue, ponceau, gentian violet, methylene blue, orange G, malachite green, and Sudan III and IV. I found that these early Grübler dyes closely resembled modern day counterparts; however, the use of synonyms was confusing and some of the fat stains were mislabeled by modern criteria. The chromatograms of some dyes exhibited smearing, probably representing multiple closely related dye species. The study of old dyes provides interesting comparisons with modern counterparts as the center of dye manufacturing is moving from Europe and the United States to Asia.     

Comparison of historic Grübler dyes with modern counterparts using thin layer chromatography.

The aniline dye industry was created in 1856 when William Perkin prepared the dye, mauve, from coal tar. Following that discovery, several dye manufacturing businesses were formed in Western Europe, most successfully in Germany. It was to these companies that early investigators turned to obtain these new dyes for the developing field of biology. In 1880, Dr. Georg Grübler started a company in Germany to supply the needs of biologists. Grübler dyes developed a reputation for excellence. In the study reported here, 29 samples of 12 Grübler dyes were compared to modern counterparts using thin layer chromatography. The dyes studied were basic fuchsine, acid fuchsine, safranine, pyronine, aniline blue, ponceau, gentian violet, methylene blue, orange G, malachite green, and Sudan III and IV. I found that these early Grübler dyes closely resembled modern day counterparts; however, the use of synonyms was confusing and some of the fat stains were mislabeled by modern criteria. The chromatograms of some dyes exhibited smearing, probably representing multiple closely related dye species. The study of old dyes provides interesting comparisons with modern counterparts as the center of dye manufacturing is moving from Europe and the United States to Asia.     

The significance of azo-reduction in the mutagenesis and carcinogenesis of azo dyes

Azo dyes are widely used in textile, printing, cosmetic, drug and food-processing industries. They are also used extensively in laboratories as either biological stains or pH indicators. The extent of such use is related to the degree of industrialization. Since intestinal cancer is more common in highly industrialized countries, a possible connection may exist between the increase in the number of cancer cases and the use of azo dyes. Azo dyes can be reduced to aromatic amines by the intestinal microflora. The mutagenicity of a number of azo dyes is reviewed in this paper. They include Trypan Blue, Ponceau 3R, Pinceau 2R, Methyl Red, Methyl Yellow, Methyl Orange, Lithol Red, Orange I, Orange II, 4-Phenylazo-Naphthylamine, Sudan I, Sudan IV, Acid Alizarin Violet N, Fast Garnet GBC, Allura Red, Ponceau SX, Sunset Yellow, Tartrazine, Citrus Red No. 2, Orange B, Yellow AB, Carmoisine, Mercury Orange, Ponceau S, Versatint Blue, Phenylazophenol, Evan's Blue and their degraded aromatic amines. The significance of azo reduction in the mutagenesis and carcinogenesis of azo dyes is discussed.     

Investigation of photosensitizing dyes for pathogen reduction in red cell suspensions.

Despite recent advances in blood safety by careful donor selection and implementation of infectious disease testing, transmission of viruses, bacteria and parasites by transfusion can still rarely occur. One approach to reduce the residual risk from currently tested pathogens and to protect against the emergence of new ones is to investigate methods for pathogen inactivation. The use of photosensitizing dyes for pathogen inactivation has been studied in both red cell and platelet blood components. Optimal properties of sensitizing dyes for use in red cell suspensions include selection of dyes that traverse cell and viral membranes, bind to nucleic acids, absorb light in the red region of the spectrum, inactivate a wide range of pathogens, produce little red cell photodamage from dye not bound to nucleic acid and do not hemolyze red cells in the dark. Early research at the American Red Cross focused on the use of a class of dyes with rigid structures, such as the phenothiazine dyes, beginning with the prototypical sensitizer methylene blue. Results revealed that methylene blue phototreatment could inactivate extracellular virus, but resulted in undesirable defects in the red cell membrane that resulted in enhanced hemolysis that became evident during extended refrigerated blood storage. In addition, methylene blue phototreatment could neither inactivate intracellular viruses nor appreciably inactivate bacteria under conditions of extracellualar viral killing. Attempts to improve intracellular viral inactivation led to the investigations of more hydrophobic phenothiazines, such as methylene violet or dimethylmethylene blue. Although these dyes could inactivate intracellular virus, problems with increased red cell membrane damage and hemolysis persisted or increased. Further studies using red cell additive storage solutions containing high levels of the impermeable ion, citrate, to protect against colloidal osmotic hemolysis as well as competitive inhibitors to limit sensitizer binding to red cell membranes revealed that photoinduced hemolysis stemmed from dye bound to the red cell membrane as well as dye free in solution. Use of red cell additive solutions to prevent colloidal-osmotic hemolysis and use of novel flexible dyes that only act as sensitizers when bound to their targets are two techniques that currently are under investigation for reducing red cell damage. Ultimately, the decision to implement a photodynamic method for pathogen reduction will be determined by weighing the risks of unintended adverse consequences of the procedure itself, such as the potential for genotoxicity and allergic reactions, against the cost and benefits of its implementation.     

Investigation of photosensitizing dyes for pathogen reduction in red cell suspensions

Despite recent advances in blood safety by careful donor selection and implementation of infectious disease testing, transmission of viruses, bacteria and parasites by transfusion can still rarely occur. One approach to reduce the residual risk from currently tested pathogens and to protect against the emergence of new ones is to investigate methods for pathogen inactivation. The use of photosensitizing dyes for pathogen inactivation has been studied in both red cell and platelet blood components. Optimal properties of sensitizing dyes for use in red cell suspensions include selection of dyes that traverse cell and viral membranes, bind to nucleic acids, absorb light in the red region of the spectrum, inactivate a wide range of pathogens, produce little red cell photodamage from dye not bound to nucleic acid and do not hemolyze red cells in the dark. Early research at the American Red Cross focused on the use of a class of dyes with rigid structures, such as the phenothiazine dyes, beginning with the prototypical sensitizer methylene blue. Results revealed that methylene blue phototreatment could inactivate extracellular virus, but resulted in undesirable defects in the red cell membrane that resulted in enhanced hemolysis that became evident during extended refrigerated blood storage. In addition, methylene blue phototreatment could neither inactivate intracellular viruses nor appreciably inactivate bacteria under conditions of extracellualar viral killing. Attempts to improve intracellular viral inactivation led to the investigations of more hydrophobic phenothiazines, such as methylene violet or dimethylmethylene blue. Although these dyes could inactivate intracellular virus, problems with increased red cell membrane damage and hemolysis persisted or increased. Further studies using red cell additive storage solutions containing high levels of the impermeable ion, citrate, to protect against colloidal osmotic hemolysis as well as competitive inhibitors to limit sensitizer binding to red cell membranes revealed that photoinduced hemolysis stemmed from dye bound to the red cell membrane as well as dye free in solution. Use of red cell additive solutions to prevent colloidal-osmotic hemolysis and use of novel flexible dyes that only act as sensitizers when bound to their targets are two techniques that currently are under investigation for reducing red cell damage. Ultimately, the decision to implement a photodynamic method for pathogen reduction will be determined by weighing the risks of unintended adverse consequences of the procedure itself, such as the potential for genotoxicity and allergic reactions, against the cost and benefits of its implementation.     

Investigation of photosensitizing dyes for pathogen reduction in red cell suspensions

Despite recent advances in blood safety by careful donor selection and implementation of infectious disease testing, transmission of viruses, bacteria and parasites by transfusion can still rarely occur. One approach to reduce the residual risk from currently tested pathogens and to protect against the emergence of new ones is to investigate methods for pathogen inactivation. The use of photosensitizing dyes for pathogen inactivation has been studied in both red cell and platelet blood components. Optimal properties of sensitizing dyes for use in red cell suspensions include selection of dyes that traverse cell and viral membranes, bind to nucleic acids, absorb light in the red region of the spectrum, inactivate a wide range of pathogens, produce little red cell photodamage from dye not bound to nucleic acid and do not hemolyze red cells in the dark. Early research at the American Red Cross focused on the use of a class of dyes with rigid structures, such as the phenothiazine dyes, beginning with the prototypical sensitizer methylene blue. Results revealed that methylene blue phototreatment could inactivate extracellular virus, but resulted in undesirable defects in the red cell membrane that resulted in enhanced hemolysis that became evident during extended refrigerated blood storage. In addition, methylene blue phototreatment could neither inactivate intracellular viruses nor appreciably inactivate bacteria under conditions of extracellualar viral killing. Attempts to improve intracellular viral inactivation led to the investigations of more hydrophobic phenothiazines, such as methylene violet or dimethylmethylene blue. Although these dyes could inactivate intracellular virus, problems with increased red cell membrane damage and hemolysis persisted or increased. Further studies using red cell additive storage solutions containing high levels of the impermeable ion, citrate, to protect against colloidal osmotic hemolysis as well as competitive inhibitors to limit sensitizer binding to red cell membranes revealed that photoinduced hemolysis stemmed from dye bound to the red cell membrane as well as dye free in solution. Use of red cell additive solutions to prevent colloidal-osmotic hemolysis and use of novel flexible dyes that only act as sensitizers when bound to their targets are two techniques that currently are under investigation for reducing red cell damage. Ultimately, the decision to implement a photodynamic method for pathogen reduction will be determined by weighing the risks of unintended adverse consequences of the procedure itself, such as the potential for genotoxicity and allergic reactions, against the cost and benefits of its implementation.     

Dye removal by immobilised fungi

Dyes are widely used within the food, pharmaceutical, cosmetic, printing, textile and leather industries. This has resulted in the discharge of highly coloured effluents that affect water transparency and gas solubility in water bodies. Furthermore, they pose a problem because of their carcinogenicity and toxicity. Therefore, removal of such dyes before discharging them into natural water streams is essential. For this, appropriate treatment technologies are required. The treatment of recalcitrant and toxic dyes with traditional technologies is not always effective or may not be environmentally friendly. This has impelled the search for alternative technologies such as biodegradation with fungi. In particular, ligninolytic fungi and their non-specific oxidative enzymes have been reported to be responsible for the decolouration of different synthetic dyes. Thus, the use of such fungi is becoming a promising alternative to replace or complement the current technologies for dye removal. Processes using immobilised growing cells seem to be more promising than those with free cells, since the immobilisation allows using the microbial cells repeatedly and continuously. This paper reviews the application of fungal immobilisation to dye removal.     

Historic dyes and how to identify them

The origin, production, and means of confirming the authenticity of some historic dyes are described. The underlying chemistry is revealed. The evolution of analytical techniques from early times, when the main criterion for a good textile dye was fitness for use, to more modern chromatographic techniques, where the emphasis is on chemical identity, is illustrated. Recent developments have led to smaller sample size requirements, greater speed of analysis and have led to a more detailed knowledge of the chemical components of historic dyes.     

Structure-photodynamic activity relationships of substituted zinc trisulfophthalocyanines

To identify optimal features of metalated sulfophthalocyanine dyes for their use as photosensitizers in the photodynamic therapy of cancer, we synthesized a series of alkynyl-substituted trisulfonated phthalocyanines and compared their amphiphilic properties to a number of parameters related to their photodynamic potency. Varying the length of the substituted alkynyl side-chain modulates the hydrophobic/hydrophilic properties of the dyes providing a linear relationship between their n-octanol/water partition coefficients and retention times on reversed-phase HPLC. Aggregate formation of the dyes in aqueous solution increased with increasing hydrophobicity while monomer formation was favored by the addition of serum proteins or organic solvent. Trisulfonated zinc phthalocyanines bearing hexynyl and nonynyl substituents exhibited high cellular uptake with strong localization at the mitochondrial membranes, which coincided with effective photocytotoxicity toward EMT-6 murine mammary tumor cells. Further increase in the length of the alkynyl chains (dodecynyl, hexadecynyl) did not improve their phototoxicity, likely resulting from extensive aggregation of the dyes in aqueous medium and reduced cell uptake. Aggregation was evident from shifts in the electronic spectra and reduced capacity to generate singlet oxygen. When monomerized through the addition of Cremophor EL all sulfonated zinc phthalocyanines gave similar singlet oxygen yields. Accordingly, differences in the tendency of the dyes to aggregate do not appear to be a determining factor in their photodynamic potency. Our results confirm that the latter in particular relates to their amphiphilic properties, which facilitate cell uptake and intracellular localization at photosensitive sites such as the mitochondria. Combined, these factors play a significant role in the overall photodynamic potency of the dyes.     

Red laser-induced fluorescence energy transfer in an immunosystem

We describe two near-infrared fluorescent squaraine dyes (Sq635 and Sq660), their spectra, their covalent linkage to proteins, and their use as donor and acceptor, respectively, in a fluorescence resonance energy transfer (FRET) immunoassay based on the use of red lasers. The dyes show quantum yields of around 10% in the free form and up to 68% when bound to proteins. If converted into their N-hydroxysuccinimide esters, they can be linked to free amino groups of proteins. To improve water solubility, two sulfo groups were introduced. The emission spectrum of Sq635 overlaps the absorption spectrum of Sq660, a fact that makes them a useful pair of dyes for use in FRET immunoassay which is demonstrated for human serum albumin/anti-human serum albumin.     

Current trends in photodynamic therapy of neoplasms and non-neoplastic diseases

The photodynamic therapy of tumors is a modern therapeutic modality for organ-preserving treatment of oncological diseases. The method is based on selective laser irradiation of tumor tissues previously sensitized by tumorotropic dyes. During the last decades, photodynamic therapy has become worldwide known as a proper approach to the treatment of the patients with malignant tumors of various locations and a number of nontumoral diseases. The characteristics of modern photosensitizers and light sources for photodynamic therapy and their clinical applications are reviewed.     

Fluorescent carbocyanine dyes allow living neurons of identified origin to be studied in long-term cultures

A prerequisite for many studies of neurons in culture is a means of determining their original identity. We needed such a technique to study the interactions in vitro between a class of spinal cord neurons, sympathetic preganglionic neurons, and their normal target, neurons from the sympathetic chain. Here, we describe how we use two highly fluorescent carbocyanine dyes, which differ in color but are otherwise similar, to identify neurons in culture. The long carbon chain carbocyanine dyes we use are lipid-soluble and so become incorporated into the plasma membrane. Neurons can be labeled either retrogradely or during dissociation. Some of the labeled membrane gradually becomes internalized and retains its fluorescence, allowing identification of cells for several weeks in culture. These dyes do not affect the survival, development, or basic physiological properties of neurons and do not spread detectably from labeled to unlabeled neurons. It seems likely that cells become retrogradely labeled mainly by lateral diffusion of dye in the plane of the membrane. If so, carbocyanine dyes may be most useful for retrograde labeling over relatively short distances. An additional feature of carbocyanine labeling is that neuronal processes are brightly fluorescent for the first few days in culture, presumably because dye rapidly diffuses into newly inserted membrane. We have used carbocyanine dyes to identify sympathetic preganglionic neurons in culture. Our results indicate that preganglionic neurons can survive in the absence of their target cells and that several aspects of their differentiation in the absence of target appear normal.     

The History of Staining Logwood Dyes

Except for the cochineal derivatives, logwood extract was the first of the important modern stains to be employed in histology. Certain other natural dyes, such as madder and indigo, had been used earlier, but they are of little significance in discussing the history of staining, because none of them nor even alizarin, the derivative of madder, are of any appreciable significance in these days of synthetic dyes. Hematoxylin, on the other hand, still continues a very important stain, and it has played an interesting part in the history of staining.     

The History of Staining Logwood Dyes

Except for the cochineal derivatives, logwood extract was the first of the important modern stains to be employed in histology. Certain other natural dyes, such as madder and indigo, had been used earlier, but they are of little significance in discussing the history of staining, because none of them nor even alizarin, the derivative of madder, are of any appreciable significance in these days of synthetic dyes. Hematoxylin, on the other hand, still continues a very important stain, and it has played an interesting part in the history of staining.     

Comparison of historic Grübler dyes with modern counterparts.

Seventeen Grübler dyes produced in Germany between 1880 and 1939 were examined in this study. These dyes were: fuchsin-bacillus, diamond fuchsin, fuchsin S acid, rubin S, safranin O water soluble, safranin yellowish water soluble, methyl eosin, Sudan III, scarlet R, auramine, orange G, aniline blue, pyronin, carmine, lithium carmine, hematein and aurantia. Spectrophotometry and staining characteristics were used to determine the maximum absorbance and efficacy of each dye in common staining techniques. The spectral curves and staining characteristics of these dyes compared well with modern dyes used as controls. Fuchsin bacillus and diamond fuchsin are synonyms for basic fuchsin. Fuchsin S acid and rubin S are synonyms for acid fuchsin. The scarlet R sample was the same as the Sudan III. The two safranins were the same. The basic fuchsin samples were unsuitable for preparation of Schiff's reagent. Both basic fuchsin and pyronin samples were less concentrated than modern counterparts. It is noteworthy that the dyes worked well after up to 100 years in storage, and this observation indicates that dyes can have a long shelf life when stored in cool, dry, air-tight conditions.     

Biodecolorization of azo,anthraquinonic and triphenylmethane dyes by white-rot fungi and a laccase-secreting engineered strain

One laccase-secreting engineered strain and four white-rot fungi were tested for their capacity to decolorize nine dyes that could be classified as azo, anthraquinonic and triphenylmethane dyes. Trametes versicolor was the most efficient of the tested strains under these experimental conditions. Anthraquinonic dyes were decolorized more easily than the other two types. Small structural differences among the dyes could significantly affect decolorization. None of the strains showed lignin peroxidase or veratryl alcohol oxidase activity. None of the dyes were decolorized completely by laccase alone. It is likely that other phenoloxidases, such as Mn-dependent and versatile peroxidase, were also involved in decolorization of the dyes.     

Remediation of dyes in textile effluent: a critical review on current treatment technologies with a proposed alternative

The control of water pollution has become of increasing importance in recent years. The release of dyes into the environment constitutes only a small proportion of water pollution, but dyes are visible in small quantities due to their brilliance. Tightening government legislation is forcing textile industries to treat their waste effluent to an increasingly high standard. Currently, removal of dyes from effluents is by physio-chemical means. Such methods are often very costly and although the dyes are removed, accumulation of concentrated sludge creates a disposal problem. There is a need to find alternative treatments that are effective in removing dyes from large volumes of effluents and are low in cost, such as biological or combination systems. This article reviews the current available technologies and suggests an effective, cheaper alternative for dye removal and decolourisation applicable on large scale.