Absorbance, absorbtance and corneal spectral sensitivities

    Photopigment spectra are usually defined by their absorbance (or extinction, or photopigment optical density) spectra. Absorbance (Aλ) is:


where Iinc is the incident light reaching the pigment and Itrans is the light transmitted through the pigment (and Iabs = Iinc - Itrans is the light absorbed by the pigment). Absorbance spectra are typically used to define photopigment spectra because their shape, when normalized (i.e., plotted as a fraction of the maximum absorbance), is independent of pigment optical density (pigment concentration). Please note that log (absorbance) values are tabulated in this databaseplease check the notes in each case.


    In contrast, the absorbtance spectra, like the spectral sensitivity of the human subject, broadens as the optical density increases. Absorbtance (Jλ) is related to Aλ according to the following equation (see Knowles & Dartnall, 1977, p. 56):


where D is the optical density of the photopigment. (Thus, when D is low the normalized absorbance and absorbtance spectra are the same shape.) To compare an absorbance spectrum with a corneally-measured human spectral sensitivity, requires several steps and assumptions. First, we must know the effective optical density D in the intact, in situ photoreceptor, so that we can calculate the absorbtance spectra, using the third equation above. Second, we must correct for absorption by the prereceptoral filters that lie between the photoreceptor and the cornea. Other factors, however, may also play a role. For example, the structure of the photoreceptor itself causes incident light to be transmitted along the photoreceptor in patterns called waveguide modal patterns (see Fig. 6 of Enoch, 1963). The fraction of the power of each modal pattern that is transmitted inside the photoreceptor to its power outside the photoreceptor decreases with the wavelength of the incident light, so that, in principle, the structure of the photoreceptor can change its spectral sensitivity (see, for example, Enoch, 1961; Enoch & Stiles, 1961; Snyder, 1975; Horowitz, 1981). Yet, it is difficult to know precisely how waveguide factors will influence the spectral sensitivity, or how to correct for them.


Photopigment spectra

    In some species, it is possible to extract a photopigment from retinas and then to measure its absorbance spectra in solution (e.g., frog rhodopsin; Dartnall, Goodeve & Lythgoe, 1936). For the human cone pigments, however, most measurements, until very recently (see below), have been made in situ, with the photopigment still in the outer segment. A principal method has been microspectrophotometry (MSP).

    In MSP work, the spectral transmission of a small measuring beam (passing transversely through the outer segment of a single cone) is compared with that of a reference beam (passing outside the cone) to derive the absorption spectrum of the outer segment (Bowmaker et al., 1978, for example). MSP measurements, however, are subject to systematic distortions due to such wavelength-dependent factors as light scattering, absorption by substances other than visual pigment (including photo-products), and focusing errors (MacNichol et al., 1983). Many of these problems can be avoided or reduced if, instead of an absorption spectrum, an action spectrum is measured. This is accomplished in suction electrode recordings, in which a single human or primate cone outer segment is drawn inside a small glass electrode, and its electrical (current) response to lights of different wavelength recorded (for example, Baylor et al., 1984).

    Both the suction electrode and MSP measurements have employed light stimuli that were passed transversely through the outer segment, rather than axially as in normal vision. This means that at the very least the axial optical density (D) of the photopigment must be known to reconstruct the corneal spectral sensitivities (see above).

    Recently, Merbs & Nathans (1992) have produced human cone photopigments from the DNA, and have measured their difference spectra in solution. They find mean values for the wavelength of maximum absorption of 426 and 532 nm for the S- and M-cone photopigments, respectively, and 552 and 557 nm for the two polymorphic variants of the L-cone photopigment.


Baylor, D. A., Nunn, B. J. & Schnapf, J. L. (1984). The photocurrent, noise and spectral sensitivity of rods of the monkey Macaca fascicularis. Journal of Physiology, 357, 575-607.

Bowmaker, J. K., Dartnall, H. J. A., Lythgoe, J. N. & Mollon, J. D. (1978). The visual pigments of rods and cones in the rhesus monkey Macaca mulatta. Journal of Physiology (London), 274, 329-348.

Dartnall, H. J. A., Goodeve, C. F. & Lythgoe, R. J. (1936). The quantitative analysis of the photochemical bleaching of visual purple solutions in monochromatic light. Proceedings of the Royal Society of London A, 156, 158-170.

Enoch, J. M. (1961). Nature of the transmission of energy in the retinal receptors. Journal of the Optical Society of America, 51, 1122-1126.

Enoch, J. M. & Stiles, W. S. (1961). The colour change of monochromatic light with retinal angle of incidence. Optica Acta, 8, 329-358.

Enoch, J. M. (1963). Optical properties of the retinal receptors. Journal of the Optical Society of America, 53, 71-85.

Horowitz, B. R. (1981). Theoretical considerations of the retinal receptor as a waveguide. In J. M. Enoch & F. L. Tobey (Ed.), Vertebrate photoreceptor optics pp. 217-300). Berlin, Heidelburg, New York: Springer-Verlag.

Knowles, A. & Dartnall, H. J. A. (1977). The photobiology of vision. In H. Davson (Ed.), The Eye, Vol 2B London/New York: Academic.

MacNichol, E. F., Levine, J. S., Mansfield, R. J. W., Lipetz, L. E. & Collins, B. A. (1983). Microspectrophotometry of visual pigments in primate photoreceptors. In J. D. Mollon & L. T. Sharpe (Ed.), Colour Vision: Physiology and Psychophysics pp. 13-38). London: Academic Press.

Merbs, S. L. & Nathans J. Absorption spectra of human cone pigments. Nature, 356, 433-435.

Snyder, A. W. (1975). Photoreceptor optics - theoretical principles. In A. W. Snyder, & R. Menzel (Ed.), Photoreceptor optics pp. 38-55). Berlin, Heidelburg, New York: Springer-Verlag.