The effect of sodium bisulfite on the removal of drugs and their metabolites interfering with the Porter-Silber reaction in the determination of urinary 17-hydroxycorticosteroids.

In the determination of urinary 17-hydroxycorticosteroids, the urine is saturated with NaHSO3 after hydrolysis with beta-glucuronidase and then extracted with methylene chloride. Sodium bisfulfite removes almost all non-steroidal impurities in the urines from patients medicated with acetylspiramycin, leucomycin, erythromycin, triacetylolenadomycin, rifampicin and tranquilizers such as chlorpromazine, which interfere with the absorption at 410 nm. in the subsquent Porter-Silber reaction. In order to increase the specificity of a routine method, a procedure conducted by Allen has been often employed: The sum of 370- and 450-nm. absorbances is subtracted from twice absorbance at 410 nm. However, the procedure could not be used in the medicated urines mentioned above, because the spectral absorption curve of these drugs and their metabolites in the Porter-Silber reaction was not a straight but a strongly convex or concave line in the 370-450-nm. range. Using the present method, interference with the Porter-Silber reaction was not found in the urines from patients medicated with chloramphenicol, minocycline, chlordiazepoxide, meprobamate, methyprylon, nitrazepam, synthetic penicillins such as hetacillin, oxacillin and cloxacillin, or cephalosporins such as cephalexin and cephalothin. However, to obtain correct values in urines from patients medicated with spironolactone, it was necessary to subject the urines to treatment with methylene chloride before enzyme hydrolysis.     

Spectral correlates of a quasi-stable depolarization in barnacle photoreceptor following red light.

1. Illumination of B. eburneus photoreceptors with intense red light produces a membrane depolarization that persists in darkness. This quasistable depolarization (latch-up) can be terminated with green light. The phenomenon was investigated with electrophysiological, spectrochemical, and microspectrophotometric techniques. 2. Latch-up was associated with a stable inward current in cells with the membrane potential voltage-clamped at the resting potential in darkness. The stable current could only be elicited at wave-lengths greater than 580 nm. 3. Light-induced current (LIC) was measured at various wave-lengths in dark-adapted photoreceptors with the membrane voltage-clamped to the resting potential. The minimum number of photons required to elicit a fixed amount of LIC occurred at 540 nm, indicating that the photoreceptor is maximally sensitive to this wave-length of light. The photoreceptor was also sensitive to wave-lengths in the near-U.V. region of the spectrum (380-420 nm). 4. Steady red adapting light reduced the magnitude of the LIC uniformly at all wave-lengths except in the near-U.V. region of the spectrum; sensitivity was reduced less in this region. 5. The spectrum for termination of the stable inward current following or during red light was shifted to the blue (peak about 510 nm) compared to the peak for LIC (peak about 540 nm). 6. Absorbance of single cells prepared under bright, red light decreased maximally at 480 nm following exposure to wave-lengths of light longer than 540 nm. 7. A pigment extract of 1000 barnacle ocelli prepared under dim, red light had a maximum absorbance change at 480 nm when bleached with blue-gree light. 8. There was no evidence in the latter two experiments of photointerconversion of pigments with absorbance maxima at 480 and 540 nm. Rather, the maximum absorption of the bleaching products seemed to occur at wave-lengths shorter than 420 nm. 9. Since latch-up induction occurs at wave-lengths longer than 580 nm, it may depend on the 540 pigment or on an undetected red absorbing pigment. 10. A photolabile pigment at 480 nm correlated most closely with termination of the stable inward current associated with latch-up.     

Ultraviolet absorption spectra and difference spectra of barley stripe mosaic and tobacco mosaic viruses in buffer and sodium dodecyl sulfate

Ultraviolet absorption spectra and difference spectra were determined for barley stripe mosaic virus (BSMV) virions and protein and for tobacco mosaic virus (TMV) virions and protein in buffer and in sodium dodecyl sulfate (SDS). The absorbance at 260 nm of BSMV virions disrupted in SDS was 92% of that of intact virions after correction for light scattering. The corresponding value for TMV was 76%. The decrease in A(260) for BSMV virions upon disruption in SDS was almost entirely due to loss of partial hyperchromocity of RNA in the virion. However, both the protein and the RNA of TMV were hyperchromic in the virion at 260 nm compared to their light absorption after disruption in SDS. The turbidity resulting from light scattering was not a linear function of virion concentration above 2 mg/ml.     

Photoinitiated polymerization of PEG-diacrylate with lithium phenyl-2,4,6-trimethylbenzoylphosphinate: polymerization rate and cytocompatibility

Due to mild reaction conditions and temporal and spatial control over material formation, photopolymerization has become a valuable technique for the encapsulation of living cells in three dimensional, hydrated, biomimetic materials. For such applications, 2-hydroxy-1-[4-(2-hydroxyethoxy) phenyl]-2-methyl-1-propanone (I2959) is the most commonly used photoinitiator (by virtue of its moderate water solubility), yet this initiator has an absorption spectrum that is poorly matched with wavelengths of light generally regarded as benign to living cells, limiting the rate at which it may initiate polymerization in their presence. In contrast, acylphosphine oxide photoinitiators, generally exhibit absorption spectra at wavelengths suitable for cell encapsulation, yet commercially available initiators of this class have low water solubility. Here, a water soluble lithium acylphosphinate salt is evaluated for its ability to polymerize diacrylated poly(ethylene glycol) (PEGDA) monomers rapidly into hydrogels, while maintaining high viability during direct encapsulation of cells. Through rheometric measurements, the time to reach gelation of a PEGDA solution with the phosphinate initiator is one tenth the time for that using I2959 at similar concentrations, when exposed to 365 nm light. Further, polymerization with the phosphinate initiator at 405 nm visible light exposure is achieved with low initiator concentrations and light intensities, precluded in polymerizations initiated with I2959 by its absorbance profile. When examined 24 h after encapsulation, survival rates of human neonatal fibroblasts encapsulated in hydrogels polymerized with the phosphinate initiator exceed 95%, demonstrating the cytocompatibility of this initiating system.     

Variation in the action spectrum of erythrolabe among deuteranopes.

1. Eight deuteranopes matched a mixture of a monochromatic light on the long wave side of the neutral point and a violet (450 nm) primary to a fixed white as well as a monochromatic light on the short wave side of the neutral point mixed with a red (650 nm) primary, to the same white. For lambda greater than 530 nm, the former set of matches defined the action spectrum of the long wave sensitive foveal cones, and for lambda less than 480 nm, the latter that of the short wave sensitive cones. 2. Individual differences in the former matches were approximately correlated with the respective ratio of the sensitivities of the wave-length of the anomaloscope primaries, in a way that individual differences of the latter were not. 3. Assuming that eye media differences alone account for the differences in long wave sensitive foveal action spectra, the spectral reflectivity of the foveal fundus was predicted for these deuteranopes. The prediction is inconsistent with measurement. 4. Thirteen deuteranopes matched monochromatic spectral lights with a green (535 nm) and a blue (460 nm) primary. The result were analysed by von Kries' method in which differences in matching due to differences in eye media absorption are obviated. The matches of five differed significantly from one another when so analysed. It was concluded that at least one of two action spectra of the foveal cones of every one of these five differed from that of all of the others. 5. The canon that deuteranopes accept normal colour matches was evaluated by confronting a single normal with five deuternopes in the analytical anomaloscope of Baker & Rushton, set in the mode of each of the five in turn. Obvious differences existed between this normal's matches and those of four of five deuteranopes. 6. Explanations for differences in the spectrum of erythrolabe in different deuteranopes are evaluated. The possibilities that all have the identical visual pigment but (a) in cones with different optical funnelling properties or (b) in different optical densities are considered. Preliminary results are not in agreement with the expectations of either of these ideas. 7. It is suggested that the visual pigment in the foveal long wave sensitive cones of different deuternopes (and of different normals) may have different extinction spectra. The idea is consistent with micro-spectrophotometric measurements of rhodopsin in individual rods from different frogs (Bowmaker, Loew & Leibman, 1975).     

Studies of the extraction of bilirubin from human amniotic fluid

The recovery of unconjugated bilirubin from human amniotic fluid was studied using dichloromethane, chloroform/isopropanol (3:1 vol/vol), and chloroform/ methanol (3:1 vol/vol) extraction of human amniotic fluid that had been supplemented with bilirubin at various concentrations. Results were compared with those obtained with conventional chloroform extraction. Mean recoveries were found to be only 28% for chloroform and 25% for dichloromethane. When the polarity of chloroform was increased by the addition of an alcohol, the mean recovery increased to only 40% for chloroform/isopropanol and 38% for chloroform/ methanol. These results suggest that extraction methods for determination of amniotic fluid "delta OD(450)" (visible spectrophotometric absorbance [optical density] of bilirubin at 450 nm) tend to underestimate the result when compared with the nonextraction (direct-scan) method, on which the Liley Prognostication Chart is based. This finding should be clinically significant, particularly if extraction and direct-scan methods are used to monitor the condition of the same patient.     

The yellow colour of the lens of man and other primates

1. Measurements have been made of the absorption spectra of the lenses of human, baboon (Papio), rhesus monkey (Macaca), squirrel monkey (Saimiri sciureus) and bush baby (Galago crassicaudatus).2. In all these species an absorption maximum was found between 365 and 368 nm.3. The pigment responsible for this absorption was water-soluble and aqueous extracts were examined. Protein-free aqueous extracts showed an additional maximum at 260 nm which could be only partially accounted for by the presence of ascorbic acid.4. Chromatography of the protein-free solution from human lenses yielded a fast-moving yellow component with a blue fluorescence. Its absorption spectrum was very similar to that of the original protein-free solution. A fast-moving yellow component from the baboon lens had a yellow fluorescence.5. The human lenses appeared to contain more of the yellow, water-soluble pigment at birth than in adult life. The concentration remains constant during adult life.6. There is evidence for the appearance of another pigment in the human lens in later adult life. It is not water-soluble and has an absorption maximum at about 330 nm.     

Pigment production by Lancefield-group-B streptococci (Streptococcus agalactiae)

Cells of group-B streptococci harvested in the late exponential phase of growth and suspended in starch-glucose phosphate-buffered saline extractor solution were observed to form and release pigment into solution. Filtrates of these solutions were analysed spectrophotometrically and two varieties of pigment were detected. Pigment, when freshly produced or in the presence of starch, had four absorption peaks at 520, 485, 455 and 435 nm. If albumin was substituted for starch in the extractor solution or if the starch-pigment complex was disrupted by treatment with amylase or by boiling, the four-peak pigment rapidly and irreversibly degraded to a second type with a single absorption band at 415 nm. The pigments formed by washed cell suspensions had absorption spectra identical to those produced by pigment formed during growth in Todd-Hewitt Broth. The formation and release of soluble pigment appeared to be an active metabolic process; a carrier molecule and an energy source were both required. Pigment yields were increased when the pH of the extractor solution was in the range 7.0-7.4 and when Mg2+, but not other divalent cations, was present. No differences in yield or type of pigment were observed when pigment was formed in anaerobic conditions. These findings support an earlier observation that group-B streptococcal pigment resembles a beta carotenoid. There is some added support for the suggestion that haemolysin and pigment production by these organisms are closely linked characteristics.     

A portable scanning reflectance spectrophotometer using visible wavelengths for the rapid measurement of skin pigments

A portable rapid scan reflectance spectrometer (400-700 nm in 2.8 s) has been developed for the measurement of cutaneous pigments. The instrument incorporates a tungsten halogen lamp light source, light transmission by fibre optics and wavelength selection by a circular variable wavelength interference filter. A microcomputer controls the instrument and processes the data. The performance of the instrument was evaluated by undertaking in vitro measurements of the reflectance spectra of blood. An index of the haemoglobin content of the sample based on the gradients of the log inverse reflectance spectrum between isobestic points at 527.5, 544 and 573 nm was devised and shown to be independent of the oxygenation of the haemoglobin. The haemoglobin index was combined with measurements at 558.5 nm, a wavelength at which absorbance is sensitive to the oxygenation of haemoglobin, to give a measure of oxygen saturation. The parameter was validated by determining the oxygen dissociation curve of red cells in plasma in vitro at pH 7.33, 37 degrees C and under a partial pressure of 40 mmHg of CO2.     

9,10‐Dithio/oxo‐Anthracene as a Novel Photosensitizer for Photoinitiator Systems in Photoresists

Five novel 9,10‐dithio/oxo‐anthracenes (ANs) are synthesized, which can be used as efficient photosensitizers for a biimidazole (BCIM) photoinitiator system in dry film photoresists. The potential of ANs as photosensitizer for BCIM is investigated in detail through fluorescence spectra, ESR, and real‐time FT‐IR. The resulting ANs possess strong absorption in the near UV–vis range from 350 to 450 nm with high molar extinction coefficients. The thiol ether and phenyl groups can make UV–vis absorption to be enhanced and red‐shifted significantly. Upon irradiation with 365 or 450 nm of light, BCIM can be photosensitized by ANs and can initiate photopolymerization of acrylate monomer (HDDA) efficiently. The dry film photoresist containing AN@BCIM photoinitiator system can form fine micro‐patterns with high resolution through 405 nm laser direct‐writing photolithography.     

Visible absorption properties of radiation exposed XR type-T radiochromic film

The visible absorption spectra of Gafchromic XR type-T radiochromic film have been investigated to analyse the dosimetry characteristics of the film with visible light densitometers. Common densitometers can use photospectrometry, fluorescent light (broad-band visible), helium neon (632 nm), light emitting diode (LED) or other specific bandwidth spectra. The visible absorption spectra of this film when exposed to photon radiation show peaks at 676 nm and 618 nm at 2 Gy absorbed doses which shift to slightly lower wavelengths (662 nm and 612 nm at 8 Gy absorbed dose) at higher doses. This is similar to previous models of Gafchromic film such as MD-55-2 and HS but XR type-T also includes a large absorption at lower visible wavelengths due to 'yellow' dyes placed within the film to aid with visible recognition of the film exposure level. The yellow dye band pass is produced at approximately 520 nm to 550 nm and absorbs wavelengths lower than this value within the visible spectrum. This accounts for the colour change from yellow to brown through the added absorption in the red wavelengths with radiation exposure. The film produces a relatively high dose sensitivity with up to 0.25 OD units per Gy change at 672 nm at 100 kVp x-ray energy. Variations in dose sensitivity can be achieved by varying wavelength analysis.     

Synthesis and Characterization of Poly(3‐alkylthiophene)s for Light‐Emitting Diodes

A serial of poly(3‐alkylthiophene)s with different alkyl chains substituted at 3‐position (3‐C_n‐PTs) was synthesized. The long‐chain alkyl group would enhance not only the processability but also the luminescence efficiency. The synthesized polymers were highly soluble in common organic solvents. The weight‐average molecular weights of the resulting polymers were in the range of 7 900–100 300 g/mol with molecular polydispersity indexes in the range of 1.88–8.70. The decomposition temperatures were in the range of 357.8–450.6°C determined by thermogravimetry analysis. A quasi‐reversible and stable electrochemical behavior for these polymers was observed in the oxidation process. The highest occupied molecular orbital, lowest unoccupied molecular orbital energy levels and band gaps were estimated by electrochemical as well as optical measurements. The UV‐Vis absorption spectra of the present polymers showed absorption bands around 400–427.5 nm in solution and 430–490 nm as thin solid films. The photoluminescence spectra showed strong light emission at ˜520 nm in chloroform and at ˜560–580 nm as thin solid film. Yellow‐orange light emission from the light‐emitting diodes with the structure of Indium‐tin oxide/poly(N‐vinylcarbazole)/3‐C_n‐PTs/Al using the polymers as the active emissive layers was achieved.     

Absorption spectroscopy of EBT model GAFCHROMIC film

The introduction of radiochromic films has solved some of the problems associated with conventional 2D radiation detectors. Their high spatial resolution, low energy dependence, and near-tissue equivalence make them ideal for measurement of dose distributions in radiation fields with high dose gradients. Precise knowledge of the absorption spectra of these detectors can help to develop more suitable optical densitometers and potentially extend the use of these films to other areas such as the measurement of the radiation beam spectral information. The goal of this study is to present results of absorption spectra measurements for the new GAFCHROMIC film, EBT type, exposed to 6 MV photon beam in the dose range from 0 to 6 Gy. Spectroscopic analysis reveals that in addition to the two main absorption peaks, centered at around 583 and 635 nm, the absorption spectrum in the spectral range from 350 to 800 nm contains six more absorption bands. Comparison of the absorption spectra reveals that previous HD-810, MD-55, as well as HS GAFCHROMIC film models, have nearly the same sensitive layer base material, whereas the new EBT model, GAFCHROMIC film has a different composition of its sensitive layer. We have found that the two most prominent absorption bands in EBT model radiochromic film do not change their central wavelength position with change in a dose deposited to the film samples.     

Optical characterization of a radiochromic film by total reflectance and transmittance measurements

The GafChromic film (GCF) MD-55-2, a radiochromic material, was examined for its optical properties through total reflectance and transmittance measurements in visible spectrum (400-700 nm). By using a multilayer model of the film and Kubelka-Munk's (KM) theory, absorption and scattering coefficients of the film sensitive layer (K and S, respectively) were obtained from measurements of irradiated and nonirradiated slides. This has allowed calculation of the absorbance A(KM) of the sensitive layer of the GCF. The model easily splits scattering from absorption. Unlike absorption, scattering is essentially insensitive to irradiation dose and decreases slowly as the wavelength increases. The scattering effect is predominant over absorption in the 400-500 nm range, while beyond 600 nm absorption prevails. The A(KM) absorbance of the sensitive layer was calculated using the K coefficient and compared with the optical densities (OD) measured considering only ballistic photons (as in a standard spectrophotometer) as well as the optical densities measured collecting all the transmitted photons (as in many densitometers). The values of A(KM) found were always lower than OD measured by the other methods and they had the best linearity on the whole visible range. These data support the hypothesis that the sensitive layer reacts to irradiation more linearly than that shown by measurements using standard commercial devices. However, in the 600-680 nm range, correction is not very important because absorption is predominant over scattering. When GCF is used for imaging, scattering produces a loss of spatial information. Consequently, it is necessary to collect only ballistic photons and to correct absorbance by K and S coefficients.     

An accurate method of plasma volume measurement by direct analysis of Evans blue spectra in plasma without dye extraction: origins of albu min-space variations during maximal exercise

The Evens blue dye (EBD) dilution method, including a dye extraction step, is a standard way of measuring plasma volume. This report describes a new direct spectrophotometric method, which is simple and specific and avoids the dye extraction step. To begin with, all the contaminants which may appear during a plasma volume study were added to plasma samples, prior to the absorbance measurements. In this way we calculated correction factors for the haemolysis and the turbidity of the plasma samples, without dye. The study of the visible spectra showed that the correction factors could be obtained by measuring absorbances at only four visible wavelengths: 780, 720, 619 and 578 nm. The addition of various amounts of contaminants to dye-containing plasma samples allowed us to obtain precise values for the absorbance errors. The previously defined correction factors were then applied, and the residual absorbance errors were found to become nil. This spectrophotometric method can be used to check the efficiency of a dye-extraction procedure, as well as to study other biological fluids containing EBD. When used to analyse a chronological series of blood samples this method appeared to provide an effective way of simultaneously studying the processes of plasma volume concentration and albumin extravasation induced by maximal exercise.     

Cell viability in optical tweezers: high power red laser diode versus Nd:YAG laser

Viability of cultivated Chinese hamster ovary cells in optical tweezers was measured after exposure to various light doses of red high power laser diodes (lambda = 670-680 nm) and a Nd:yttrium-aluminum-garnet laser (lambda = 1064 nm). When using a radiant exposure of 2.4 GJ/cm2, a reduction of colony formation up to a factor 2 (670-680 nm) or 1.6 (1064 nm) as well as a delay of cell growth were detected in comparison with nonirradiated controls. In contrast, no cell damage was found at an exposure of 340 MJ/cm2 for both wavelengths, and virtually no lethal damage at 1 GJ/cm2 applied at 1064 nm. Cell viabilities were correlated with fluorescence excitation spectra and with literature data of wavelength dependent cloning efficiencies. Fluorescence excitation maxima of the coenzymes NAD(P)H and flavins were detected at 365 and 450 nm, respectively. This is half of the wavelengths of the maxima of cell inactivation, suggesting that two-photon absorption by these coenzymes may contribute to cellular damage. Two-photon excitation of NAD(P)H and flavins may also affect cell viability after exposure to 670-680 nm, whereas one-photon excitation of water molecules seems to limit cell viability at 1064 nm.     

In vitro determination of normal and neoplastic human brain tissue optical properties using inverse adding-doubling

To complement a project towards the development of real-time optical biopsy for brain tissue discrimination and surgical resection guidance, the optical properties of various brain tissues were measured in vitro and correlated to features within clinical diffuse reflectance tissue spectra measured in vivo. Reflectance and transmission spectra of in vitro brain tissue samples were measured with a single-integrating-sphere spectrometer for wavelengths 400-1300 nm and converted to absorption and reduced scattering spectra using an inverse adding-doubling technique. Optical property spectra were classified as deriving from white matter, grey matter or glioma tissue according to histopathologic diagnosis, and mean absorption and reduced scattering spectra were calculated for the three tissue categories. Absolute reduced scattering and absorption values and their relative differences between histopathological groups agreed with previously reported results with the exception that absorption coefficients were often overestimated, most likely due to biologic variability or unaccounted light loss during reflectance/transmission measurement. Absorption spectra for the three tissue classes were dominated by haemoglobin absorption below 600 nm and water absorption above 900 nm and generally determined the shape of corresponding clinical diffuse reflectance spectra. Reduced scattering spectral shapes followed the power curve predicted by the Rayleigh limit of Mie scattering theory. While tissue absorption governed the shape of clinical diffuse reflectance spectra, reduced scattering determined their relative emission intensities between the three tissue categories.     

Visible dye light absorption properties of processed radiographic film

The visible absorption spectra of Kodak X-Omat V film, which had been exposed to various doses of radiation, have been investigated to analyse the dosimetry characteristics of the film with various densitometers. Common densitometers can use fluorescent light (broad band visible), helium-neon (632 nm) or other spectra of specific bandwidth. The visible absorption spectra show a slight peak in absorption at approximately 580 nm and another at 630 nm caused by the base material of the film. The optical density of the film is shown to increase almost equally at all wavelengths within the visible region with increases in applied dose. By evaluating the results for the broad band spectra and specific wavelength optical density it is shown that a relatively uniform response is expected for all densitometers that work within the visible region as well as in selected infrared wavelengths. Thus similar optical density to dose response curves for X-Omat V radiographic film should be produced for all types of densitometers, no matter what type of light source is used for illumination. Thus it is most efficient to have a densitometer with a light source suitable for radiochromic film, which can also be used with radiographic film.     

Simulation study of in vitro glucose measurement by NIR spectroscopy and a method of error reduction

The effects of some important factors on the blood glucose measurements by NIR spectroscopy are investigated by numerical simulation, and a method is proposed to significantly reduce the prediction errors induced by these effects. The changes in the absorbance spectra with the changes in the glucose concentration, temperature and scattering characteristics of background tissue are obtained by a Monte Carlo simulation of light propagation for the wavelength range from 1200 nm to 1800 nm. The glucose concentration is predicted by applying a multivariate analysis to the numerically simulated spectra. This process estimates the errors in the prediction of the glucose concentration induced by the temperature and scattering changes. It has been found that only 1 C change in the temperature or only 1% change in the scattering coefficient induces about 500 mg dl(-1) or 300 mg dl(-1) errors, respectively, in the prediction of the glucose concentration. These errors can be significantly reduced to less than 20 mg dl(-1) of the glucose concentration by incorporating the effects of the temperature and scattering characteristics on the spectra to the multivariate analysis.     

An attempt to analyse colour reception by electrophysiology

1. The problem of colour reception is that we do not know the action spectra of the visual pigments involved, the nature of the signals generated nor the interaction between these signals. We only know the incident light and the electric results of interaction.2. In Part 1 we show that S-potentials from red/green (R/G) units saturated with deep red light show this property: added green light pulls down the ceiling of depolarization, but more added red had no power to raise it again. Thus lights that depress the deep red ceiling equally stimulate the green pigment equally. From this the action spectrum of the green pigment can be obtained.3. If we assume that only two visual pigments are involved in the R/G unit, and that lights which do not pull down the deep red ceiling are below the threshold for green cones, then in this range only the red pigment is excited and we may obtain its action spectrum. Its maximum is at 680 nm where no visual pigment so far has been found.4. In Part 2 we consider the following mathematical problem: ;Is it possible that two pigments of given action spectra could combine their outputs in such a way that the resultant would be identical with the output of a third pigment of given action spectrum, for every intensity of every monochromatic light?' The solution shows that this is always mathematically possible, and the necessary interaction function is deduced.5. It is shown further that if the log action spectra are the ;visual parabolas' that resemble Dartnall's nomogram, then the interaction function is simply a linear transform such as Hartline & Ratliff (1957) have found with lateral inhibition in Limulus and Donner & Rushton (1959) with silent substitution in the frog.6. An interaction that matches a single pigment to perfection for all monochromatic lights will not match it for certain mixtures. By this criterion the 680 nm excitability is a pigment and not the resultant of two other pigments, i.e. pigments more excitable in other spectral regions.7. In Part 3 monochromatic lights are matched by red+green mixtures that give identical responses. From this the action spectrum of the red pigment may be obtained without involving nerve organization (except as a null detector). The result, which has one arbitrary constant, is given by the curves of Fig. 10, the continuous curve R or one of the dotted curves. Of these only curve R is acceptable.8. Knowing the action spectra for red and green cones we may consider what signals are generated and how they interact to give the records. Figure 11 suggests a model that will account for the size and sign of S-potentials as function of the quantum catch by the two pigments. It does not embrace the time or space parameters which can be very complex.