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We describe herein a series of new derivatives of tetrachloro tetraiodo (rose bengal) and report their emission and absorption characteristics. Photochemists and photobiologists may find several of these derivatives useful as alternative sources of singlet oxygen.
Altogether 117 patients with cervical cancer on combined radiation therapy were examined. They were examined before the start of radiation therapy, after a focal dose of 35-40 Gy, immediately after the termination of irradiation and in 3-12 mos. after treatment. Using a method of dynamic computerized scintigraphy with 131I-Bengal rose absorptive-excretory function of the liver was studied; the level of ferritin was determined too. Combined radiation therapy was shown to cause hepatic disorders that manifest themselves in the suppression of absorptive-excretory function of the liver and a decreased level of ferritin. The most noticeable changes were recorded in the patients examined immediately after the termination of irradiation. Results of both methods show good correlation.
The photooxidation of N,N-diethylhydroxylamine (DEHA) by Rose Bengal (RB) has been investigated in micellar and nonmicellar aqueous solutions. We measured the quantum yield of oxygen consumption forming H2O2 and monitored two intermediates, the superoxide and diethylnitroxide radicals. When the pH was varied, the quantum yield of oxidation remained constant for 6 < pH < 10.5, decreased in acidic pH, and increased considerably in NaOH solution; these changes could be attributed to the protonation and dissociation processes of the > N-OH moiety of DEHA. The formation of diethylnitroxide radical was enhanced by superoxide dismutase or strong alkaline solution. Around neutral pH, the oxidation proceeded mainly via electron transfer from DEHA to the RB triplet (kq = 10(7) M-1 s-1) with little 1O2 participation (kq < 10(5) M-1 s-1). However, when RB was incorporated into micelles in alkaline solution, the contribution of the singlet oxygen pathway increased at the expense of electron transfer, which was inhibited by the less polar micellar environment. Dark autoxidation of DEHA was accelerated by heavy metal impurities and increased very strongly in NaOH solution.
We have shown in a previous paper that irradiation of rabbit skin sites injected with Rose Bengal (RB) produces immediate increase in vascular permeability and accumulation of PMNs. Studies on the development of temporary tolerance and the biological parameters related to the development of such tolerant state by repeated exposure to light of RB-injected sites are reported here. The increase in VP and PMN migration induced by RB (10 nmol) are of an immediate nature, i.e., occur within the first 3 h of irradiation, and the reaction subsides gradually after 24 h. When such moderate insult is repeated, the skin becomes tolerant to subsequent exposure to light in the presence of RB. This tolerant state is temporary, i.e., the desensitized sites are fully recovered in 72 h. The loss of responsiveness of RB-injected sites previously exposed to light was not due to diffusion of the injected dye from the sites since reinjected sites also showed reduced response and the sites injected three days before but not irradiated showed normal response. The sites that were made tolerant to RB-induced phototoxic reactions, when injected with compound 48/80, an agent known to degranulate mast cells, did not show an increase in VP. This suggests that either the mast cells were depleted from the sites or the mast cells in the sites were rendered refractory by previous exposure to light. It was also found that the sites made tolerant to RB plus light were unresponsive to exogenously injected histamine. The sites tolerant to RB plus light when injected with zymosan-activated serum (ZAS) did not stimulate the migration of PMNs. This loss of chemotactic response to ZAS may have relevance to photodamage of vascular endothelium. These observations are discussed in relation to the development of the tolerant state by repeated exposures to subthreshold doses of light in solar urticaria.
A hospital cum community based study indicated that rose Bengal staining of the eye could not only confirm clinically diagnosed cases of conjunctival xerosis but also detect those missed by clinical examination. However, some cases thus detected did not respond to a massive oral dose of vitamin A.
The Rose-Bengal plate test (RBPT) was performed on 488 patients with flu-like symptoms from Narok district. There was poor agreement between RBPT results from four health facilities in Narok and from the central veterinary laboratory (CVL). Agreement was poorer for the three rural dispensaries than for the District Hospital. On the other hand, for tests conducted at the CVL, there was good agreement between RBPT, serum agglutination test (SAT) and complement fixation test (CFT) results, indicating that all these tests were probably performing well. Better training and quality control and the use of white rather than a clear background surface for judging agglutination results are recommended to improve the performance of test results in Narok District health facilities.
The thyrotoxic effect of Rose bengal (RB) (4,5,6,7-tetrachloro-2',4',5',7'-tetraiodofluorescein disodium salt; Food Red No. 105) was examined in male (C57BL/6N X C3H/N) F1 mice. They were given drinking-water containing RB at levels of 0 (control), 0.125 and 0.250% for 2 weeks. The effect resulted in decreases in serum levels of 3,5,3'-triiodothyronine (T3) and thyroxine (T4), and slight increases in serum 3,3',5'-triiodothyronine (rT3) levels and thyroid weight, but no difference in the values for the body-weight gain, serum thyroid stimulating hormone (TSH) levels and thyroid peroxidase (TPO) activities. However, the in vitro inhibitory effect of RB on TPO activity was observed by addition of RB to the TPO-catalyzed guaiacol oxidation. These results suggest that RB might have weak goitrogenic properties, inhibiting the peripheral conversion of T4 to T3 and/or inhibiting TPO to lead a decrease of T4 and T3 formation.
The results of a comparative study among complement fixation (CFT), plate agglutination (PAT), tube agglutination (TAT) and Rose Bengal plate tests ( RBPT ) to the serodiagnosis of brucellosis in Indian buffaloes are reported. Sera from 212 buffaloes unvaccinated against brucellosis were examined and the CFT was able to reveal significant titres in sera with low agglutinating titres. From 109 sera which did not show agglutination titres in the PAT, four showed complement fixing titre greater than 1 in 200. All the positive sera to the RBPT gave complement fixing titre equal to or greater than 1 in 20. In sera that showed negative result to the RBPT the CFT was able to reveal relatively high titres. From 131 sera negative to the RBPT five showed complement fixing titres greater than 1 in 60.
The present study was to determine serological activity of cattle anti-Brucella antibodies in rose bengal plate test with regard to classical tests, i.e. standard agglutination test (SAT) and CFT (Complement Fixation Test). The serum containing antibodies of IgM class characteristic of the early stage of infection was taken from a calf on 13th day after the administration of Br. abortus S19 killed strain. The serum containing IgG antibodies (IgG1 and IgG2 subclasses) indicating the chronic brucellosis was taken from a cow infected naturally. IgM antibodies were isolated from serum by column chromatography on Sefadeks G-200 gel and IgG1 and IgG2 by column chromatography on DEAE cellulose. It was found out that IgM, IgG1, IgG2 were active in RBPT, SAT and IgM, IgG1 in CFT. The activity of anti-Brucella immunoglobulins in RBPT proves the possibility of diagnosing the above mentioned infections using this test. Taking into account titer height, in particular tests with certain Ig class, it can be said that acidic medium in RBPT reduces not selectively the activity of all immunoglobulin classes comparing with SAT and CFT, and that RBPT is typical qualitative test more valuable than SAT. The reduction of activity of all immunoglobulin classes in RBPT at significantly lower content of M immunoglobulin than IgG1 in the serum may explain the very high specificity of the test. Moreover, it was found that half the amount of the antigen in RBPT has a significant effect on the increase of test sensitivity.
To investigate the nonlinear kinetics in the hepatobiliary transport of rose bengal (RB), the time profiles of plasma concentration and biliary excretion rate after its i.v. administration at various doses were measured in rats. The total body clearance decreased remarkably with increased dose. The hepatic uptake clearance also showed a similar dose dependency, and saturation of hepatic uptake at least partly accounts for the dose-dependent change in total body clearance. The peak biliary excretion rate approached the transport maximum (approximately 150 nmol min-1 kg-1) with increased dose. To further clarify which process in RB hepatobiliary transport has nonlinearity, we analysed thus obtained data based on a three-compartment model. The hepatic uptake and sequestration rate constants decreased remarkably with increased dose. The initial hepatic uptake rates assessed from the plasma disappearance rate during the early phase fit well to the Michaelis-Menten equation with a saturable and a nonsaturable component. The maximum uptake velocity and Michaelis constant were 4.7 mumol min-1 kg-1 and 360 microM, respectively. That hepatic uptake has a much higher capacity (about 30 fold) than biliary excretion suggests that biliary excretion can be a rate-determining process in the overall hepatobiliary transport of RB. We conclude that the saturation of both hepatic uptake and biliary excretion could be the main causes for the nonlinear pharmacokinetics of hepatobiliary transport of RB.
This paper describes a study on the sequence of the phototoxic effects of rose bengal (RB), a fluorescein derivative used as a vital stain in the diagnosis of certain external ocular diseases. Bovine melanotic RPE cells were grown in culture. These cells were labeled with [51Cr] and exposed to visible light in the presence of various concentrations of RB; the leakage of [51Cr] from the cells was used as a measure of cell lysis. Exposure to light of the cells with 0.3-10 microM RB induced approximately 13 to 43% cell lysis. The lysis progressively increased when the exposure time was varied from 10 to 30 min. A relatively short period of irradiation in the presence of RB was sufficient to produce sublytic cellular injury which could subsequently lead to complete cell lysis even in the absence of the photochemical treatment. The dark reaction was time-dependent, and reached a maximum for a given irradiation period. Our results thus show that there are two different processes that could eventually lead to the cell lysis: (a) a phototoxic effect which caused a sublytic damage and (b) a dark reaction that followed.
The quenching rate constants of the excited triplet state of Rose Bengal (RB) by oxygen (k(obs)) were measured in ethylene glycol (EG) at different temperatures using nanosecond laser flash photolysis. Although a plot of the quenching rate constant k(obs) for RB triplet state vs oxygen concentration is linear at 20 degrees C, the oxygen dependence of k(obs) does not exhibit linearity but upward curvature at high temperatures from 130 to 140 degrees C. The upward curvature at high temperatures is not well-described by a kinetic scheme first postulated by Gijzeman et al., which is characterized by exciplex formation and a unimolecular dissociation of the exciplex to products, but instead by a more comprehensive mechanism involving a bimolecular dissociation in addition to a unimolecular one. The measurements of the oxygen dependence of k(obs) for RB triplet state at different temperatures yielded a reaction enthalpy for the exciplex formation of 150 kJ mol(-1). Due to the large exothermic reaction enthalpy, equilibrium was obtained for the exciplex at 20 degrees C even at low oxygen concentration and the bimolecular quenching by oxygen became the major dissociation process. The equilibrium attainment and bimolecular dissociation provide a linear oxygen dependence of k(obs) to all outward appearances. Therefore, linearity does not always mean that exciplex dissociation proceeds solely through a unimolecular mechanism.
In Machakos Hospital, Kenya, the sera of 452 patients suspected of brucellosis were examined with the Rose Bengal Plate Test (RBPT), the complement fixation test (CFT) and the tube serum agglutination test (SAT). The percentage of false positive findings on the RBPT was 0.22% when compared with the SAT, 0.44% with the CFT. The percentage of false negative findings on the RBPT was 3% when compared with the SAT, 0.88% with the CFT. A good correlation between the RBPT and both the SAT and the CFT was also found by Morgan et al. (1969). The RBPT may be considered as a highly reliable test in the diagnosis of human brucellosis. The test is especially valuable in areas where laboratory facilities are not available and can be used as a screening test in areas with good laboratory facilities.
The aim of the present investigation was to elucidate whether the Golgi apparatus undergoes photodamage following administration of the fluorogenic substrates Rose Bengal acetate (RBAc) and irradiation at the appropriate wavelength. Human HeLa cells were treated in culture and the changes in the organization of the Golgi apparatus were studied using fluorescence confocal microscopy and electron microscopy, after immunocytochemical labeling. To see whether the cytoskeletal components primarily involved in vesicle traffic (i.e., microtubules) might also be affected, experiments of tubulin immunolabeling were performed. After treatment with RBAc and irradiation, cells were allowed to grow in drug-free medium for different times. 24 hr after irradiation, the cisternae of the Golgi apparatus became packed, and after 48-72 hr they appeared more fragmented and scattered throughout the cytoplasm; these changes in the organization of the Golgi cisternae were confirmed at electron microscopy. Interestingly enough, apoptosis was found to occur especially 48-72 h after irradiation, and apoptotic cells exhibited a dramatic fragmentation of the Golgi membranes. The immunolabeling with anti-tubulin antibody showed that microtubules were also affected by irradiation in RBAc-treated cells.
The present studies were undertaken to quantitate the initial inflammatory response produced by the photo-generated reactive species in rabbit skin. Rose bengal (RB), a photosensitizer dye, was injected into the skin sites at various concentrations and exposed to UV-visible light for 30-120 min. The increase in vascular permeability and the accumulation of PMNs were investigated using 125I-labeled albumin and 51Cr-labeled PMNs. RB at a concentration of 1 nmol with 120-min exposure to light enhanced vascular permeability by 3.7 times and accumulation of PMNs by 3.3 times. As low as 0.01 nmol of RB produced discernible effects. beta-Carotene (0.1 nmole) inhibited the inflammatory response by 75-100%, suggesting that the reactive species involved in this response was predominantly singlet oxygen. The increase in vascular permeability was inhibited by 48-70% by 25 micrograms of chlorpheniramine maleate. It is therefore suggested that histamine plays a major role in the initial vascular response. The studies demonstrate that this rabbit model is suitable for the quantitation of photoinduced inflammatory response which is not observable by gross anatomic procedures.
A new technique for the staining of the corneal endothelium is presented, in which the actions of the vital stain, rose bengal, and the intercellular stain, alizarin red S, are combined. The technique is an improvement over other simple evaluative methods, because it defines both healthy and damaged cells, and thereby permits an investigator to accurately quantify endothelial cell damage.
PURPOSE: In severely tear-deficient dry eye patients whose puncta had been occluded, corneal and conjunctival rose bengal (RB) staining were graded and compared to before occlusion. SUBJECTS AND METHODS: Subjects were 17 eyes of 13 severely tear-deficient dry eye patients whose puncta had been successfully occluded (corneal fluorescein staining showed A 0 D 0 after occlusion). Corneal and conjunctival RB staining were graded 0, 1, 2 or 3, according to severity [B 1] at the cornea and at the temporal and nasal bulbar conjunctiva. The scores were then compared to those before punctal occlusion. To enable comparison of pre- and post-occlusion ocular surface concentrations of RB [B 2], post-occlusion concentration was adjusted by absorbing tears from the conjunctival sac with Schirmer's test paper until the lower meniscus height was equal to that before occlusion, as monitored by video-meniscometer. RB dye was then instilled into the patients conjunctival sac. RESULTS: RB staining score at the cornea decreased significantly; in contrast, no significant improvement was found at the nasal and temporal conjunctiva (p = 0.0025, p = 0.05, p = 0.7, respectively, Wilcoxon test). CONCLUSION: In accordance with the basic interpretation of RB staining, these results imply that RB positive staining [B 3] on the conjunctival surface is caused by a factor other than aqueous deficiency.
The effects of erythrosine (Red 3), rose bengal B (Red 105) and thyroidectomy on the development of thyroid tumor were examined in male Wistar rats treated with N-bis(2-hydroxypropyl)nitrosamine (DHPN). Red 3 and Red 105 were used at 4% in the basal diet and were administered for 19 weeks from week 2 to 20. Thyroidectomy was performed by resection of the left lobe at week 4. Single injection of DHPN was performed intraperitoneally at 280 mg per 100 g body weight at the beginning of the experiment. Red 3 and Red 105 significantly promoted the development of thyroid tumors in thyroidectomized rats given DHPN, but had no significant effect in non-thyroidectomized rats. The incidence of thyroid tumors was 91% in rats with partial thyroidectomy, Red 3 and DHPN, 100% in rats with partial thyroidectomy, Red 105 and DHPN, and 64% in rats with partial thyroidectomy and DHPN. Serum TSH was 5.5 +/- 3.1 ng/ml in rats with partial thyroidectomy, Red 3 and DHPN, 2.1 +/- 2.2 ng/ml in rats with partial thyroidectomy, Red 105 and DHPN, and 1.5 +/- 0.5 ng/ml in rats with partial thyroidectomy and DHPN.