Microspectrophotometric characterization and localization of three visual pigments in the compound eye ofNotonecta glauca L. (Heteroptera)

Summary The absorption maxima ( _max) of the visual pigments in the ommatidia ofNotonecta glauca were found by measuring the difference spectra of single rhabdomeres after alternating illumination with two different adaptation wavelengths. All the peripheral rhabdomeres contain a pigment with an extinction maximum at 560 nm. This pigment is sensitive to red light up to wavelengths > 700 nm. In a given ommatidium in the dorsal region of the eye, the two central rhabdomeres both contain one of two pigments, either a pigment with an absorption maximum in the UV, at 345 nm, or — in neighboring rhabdoms — a pigment with an absorption maximum at 445 nm. In the ventral part of the eye only the pigment absorbing maximally in the UV was found in the central rhabdomeres. The spectral absorption properties of various types of screening-pigment granules were measured.     

Meta-Analysis of the Detection of Plant Pigment Concentrations Using Hyperspectral Remotely Sensed Data

Passive optical hyperspectral remote sensing of plant pigments offers potential for understanding plant ecophysiological processes across a range of spatial scales. Following a number of decades of research in this field, this paper undertakes a systematic meta-analysis of 85 articles to determine whether passive optical hyperspectral remote sensing techniques are sufficiently well developed to quantify individual plant pigments, which operational solutions are available for wider plant science and the areas which now require greater focus. The findings indicate that predictive relationships are strong for all pigments at the leaf scale but these decrease and become more variable across pigment types at the canopy and landscape scales. At leaf scale it is clear that specific sets of optimal wavelengths can be recommended for operational methodologies: total chlorophyll and chlorophyll a quantification is based on reflectance in the green (550–560nm) and red edge (680–750nm) regions; chlorophyll b on the red, (630–660nm), red edge (670–710nm) and the near-infrared (800–810nm); carotenoids on the 500–580nm region; and anthocyanins on the green (550–560nm), red edge (700–710nm) and near-infrared (780–790nm). For total chlorophyll the optimal wavelengths are valid across canopy and landscape scales and there is some evidence that the same applies for chlorophyll a.     

Phototaxis in monochromatic light and microspectrophotometry of the cerebral eye of the rotifer Brachionus calyciflorus

The observed wavelength-dependent variations in the phototaxis of the rotifer Brachionus calyciflorus inform us only partially about the spectral characteristics of the sensory pigment of the eye, since these variations are also linked to the absorption spectrum of the accessory pigment(s).

The absence of phototaxis between 420 nm and 500 nm is due to the lack of sensitivity of the sensory pigments at these wavelengths

The absence of response between 650 nm and 700 nm is due to a drop in the absorbance of the accessory pigments, which consequently no longer play a screening role at these wavelengths

The existence of oriented responses between 350 nm and 420 nm and between 500 nm and 650 nm, is due to the joint intervention of the two types of pigments at these wavelengths

     

Molecular basis of spectral tuning in the red- and green-sensitive (M/LWS) pigments in vertebrates

Vertebrate vision is mediated by five groups of visual pigments, each absorbing a specific wavelength of light between ultraviolet and red. Despite extensive mutagenesis analyses, the mechanisms by which contemporary pigments absorb variable wavelengths of light are poorly understood. We show that the molecular basis of the spectral tuning of contemporary visual pigments can be illuminated only by mutagenesis analyses using ancestral pigments. Following this new principle, we derive the "five-sites" rule that explains the absorption spectra of red and green (M/LWS) pigments that range from 510 to 560 nm. Our findings demonstrate that the evolutionary method should be used in elucidating the mechanisms of spectral tuning of four other pigment groups and, for that matter, functional differentiations of any other proteins.     

Microspectrophotometry of the dioptric apparatus and compound rhabdom of the moth (Manduca sexta) eye

Absorption spectra were obtained by microspectrophotometric (MSP) axial measurements of the compound rhabdom of the night moth Manduca sexta. Difference spectra derived from partial or complete bleaches revealed the evidence of four visual pigments with approximate λ_max at 350, 450, 490, and 530 nm. Upon bleaching with light of the pigment maximum at 21°C, pH 7·4–8·5, each pigment, save the u.v.-sensitive one, formed a photoproduct whose spectral maximum (ca. 370 nm) was indicative of a mixture of free and bound retinal. Rarely, small amounts of an additional photoproduct (λ_max 325–330 nm) formed, which is suggestive of retinol. The u.v.-sensitive pigment, when irradiated with u.v., formed an unknown photoproduct (λ_max 290–300 nm). Bleaching kinetics were of first order. Separate absorption determinations through lens or crystalline cones showed each component of the dioptric apparatus served as a filter effecting a sharp decrease in corneal transmission at 310 nm while being increasingly transparent from near u.v. to red. The survival benefits accruing to a largely nocturnal moth with a presumptive colour vision mechanism are discussed.     

Steady-state polarized light spectroscopy of isolated Photosystem I complexes

Monomeric and trimeric Photosystem I core complexes from the cyanobacterium Synechocystis PCC 6803 and LHC-I containing Photosystem I (PS I-200) complexes from spinach have been characterized by steady-state, polarized light spectroscopy at 77 K. The absorption spectra of the monomeric and trimeric core complexes from Synechocystis were remarkably similar, except for the amplitude of a spectral component at long wavelength, which was about twice as large in the trimeric complexes. This spectral component did not contribute significantly to the CD-spectrum. The (77 K) steady-state emission spectra showed prominent peaks at 724 nm (for the Synechocystis core complexes) and at 735 nm (for PS I-200). A comparison of the excitation spectra of the main emission band and the absorption spectra suggested that a significant part of the excitations do not pass the red pigments before being trapped by P-700. Polarized fluorescence excitation spectra of the monomeric and trimeric core complexes revealed a remarkably high anisotropy (0.3) above 705 nm. This suggested one or more of the following possibilities: 1) there is one red-most pigment to which all excitations are directed, 2) there are more red-most pigments but with (almost) parallel orientations, 3) there are more red-most pigments, but they are not connected by energy transfer. The high anisotropy above 705 nm of the trimeric complexes indicated that the long-wavelength pigments on different monomers are not connected by energy transfer. In contrary to the Synechocystis core complexes, the anisotropy spectrum of the LHC I containing complexes from spinach was not constant in the region of the long-wavelength pigments, and decreased significantly below 720 nm, the wavelength where the long-wavelength pigments on the core complexes start to absorb. These results suggested that in spinach the long-wavelength pigments on core and LHC-I are connected by energy transfer and have a non-parallel average Q_y(0-0) transitions.Abbreviations PS Photosystem - P Primary donor - Chl chlorophyll - LHC light-harvesting complex - CD circular dichroism - LD linear dichroism - BisTris 2-[bis(2-hydroxyethyl)amino]-2-hydroxy-methylpropane-1,3-diol - RC reaction center     

Excitation dynamics and heterogeneity of energy equilibration in the core antenna of photosystem I from the cyanobacterium Synechocystis sp. PCC 6803

Energy equilibration in the photosystem I core antenna from the cyanobacterium Synechocystis sp. PCC 6803 was studied using femtosecond transient absorption spectroscopy at 298 K. The photosystem I core particles were excited at 660, 693, and 710 nm with 150 fs spectrally narrow laser pulses (fwhm = 5 nm). Global analysis revealed three kinetic processes in the core antenna with lifetimes of 250-500 fs, 1.5-2.5 ps, and 20-30 ps. The first two components represent strongly excitation wavelength-dependent energy equilibration processes while the 20-30 ps phase reflects the trapping of energy by the reaction center. Excitation into the blue and red edge of the absorption band induces downhill and uphill energy flows, respectively, between different chlorophyll a spectral forms of the core. Excitation at 660 nm induces a 500 fs downhill equilibration process within the bulk of antenna while the selective excitation of long-wavelength-absorbing chlorophylls at 710 nm results in a 380 fs uphill energy transfer to the chlorophylls absorbing around 695-700 nm, presumably reaction center pigments. The 1.5-2.5 ps phases of downhill and uphill energy transfer are largely equivalent but opposite in direction, indicating energy equilibration between bulk antenna chlorophylls at 685 nm and spectral forms absorbing below 700 nm. Transient absorption spectra with excitation at 693 nm exhibit spectral evolution within approximately 2 ps of uphill energy transfer to major spectral forms at 680 nm and downhill energy transfer to red pigments at 705 nm. The 20-30 ps trapping component and P(700) photooxidation spectra derived from data on the 100 ps scale are largely excitation wavelength independent. An additional decay component of red pigments at 710 nm can be induced either by selective excitation of red pigments or by decreasing the temperature to 264 K. This component may represent one of the phases of energy transfer from inhomogeneously broadened red pigments to P(700). The data are discussed based on the available structural model of the photosystem I reaction center and its core antenna.     

Photoreceptors and visual pigments in three species of newts

Photoreceptor composition and retinal visual pigments in three newt (Caudata, Salamandridae, Pleurodelinae) species (Pleurodeles waltl, Lissotriton (Triturus) vulgaris, and Cynops orientalis) were studied by light microscopy and single-cell microspectrophotometry. Retinas of all three species contain “red” (rhodopsin/porphyropsin) rods, large and small single cones, and double cones. Large single cones and both components of double cones contain red-sensitive (presumably LWS) visual pigment whose absorption spectrum peaks between 593 and 611 nm. Small single cones are either blue- (SWS2, maximum absorption between 470 and 489 nm) or UV-sensitive (SWS1, maximum absorption between 340 and 359 nm). Chromophore composition of visual pigments (A₁ vs. A₂) was assessed both from template fitting of absorption spectra and by the method of selective bleaching. All pigments contained a mixture of A₁ (11-cis retinal) and A₂ (11-cis-3,4-dehydroretinal) chromophore in the proportion depending on the species and cell type. In all cases, A₂ was dominant. However, in C. orientalis rods the fraction of A₁ could reach 45%, while in P. waltl and L. vulgaris cones it did not exceed 5%. Remarkably, the absorption of the newt blue-sensitive visual pigment was shifted by up to 45 nm toward the longer wavelength, as compared with all other amphibian SWS2-pigments. We found no “green” rods typical of retinas of Anura and some Caudata (ambystomas) in the three newt species studied.     

Ultraviolet vision in birds: the importance of transparent eye media

Ultraviolet (UV)-sensitive visual pigments are widespread in the animal kingdom but many animals, for example primates, block UV light from reaching their retina by pigmented lenses. Birds have UV-sensitive (UVS) visual pigments with sensitivity maxima around 360–373 nm (UVS) or 402–426 nm (violet-sensitive, VS). We describe how these pigments are matched by the ocular media transmittance in 38 bird species. Birds with UVS pigments have ocular media that transmit more UV light (wavelength of 50% transmittance, λ_T0.5, 323 nm) than birds with VS pigments (λ_T0.5, 358 nm). Yet, visual models predict that colour discrimination in bright light is mostly dependent on the visual pigment (UVS or VS) and little on the ocular media. We hypothesize that the precise spectral tuning of the ocular media is mostly relevant for detecting weak UV signals, e.g. in dim hollow-nests of passerines and parrots. The correlation between eye size and UV transparency of the ocular media suggests little or no lens pigmentation. Therefore, only small birds gain the full advantage from shifting pigment sensitivity from VS to UVS. On the other hand, some birds with VS pigments have unexpectedly low UV transmission of the ocular media, probably because of UV blocking lens pigmentation.     

Tuning of photoreceptor spectral sensitivity in fireflies (Coleoptera: Lampyridae)

Sexual communication between male and female fireflies involves the visual detection of species-specific bioluminescent signals. Firefly species vary spectrally in both their emitted light and in the sensitivity of the eye, depending on the time when each is active. Tuning of spectral sensitivity in three firefly species that occupy different photic niches was investigated using light and electron microscopy, microspectrophotometry, and intracellular recording to characterize the location and spectral absorption of the screening pigments that filter incoming light, the visual pigments that receive this filtered light, and the visual spectral sensitivity. Twilight-active species had similar pink screening pigments, but the visual pigment of Photinus pyralis peaked near 545 nm, while that of P. scintillans had a λ_max near 557 nm. The night-active Photuris versicolor had a yellow screening pigment that was uniquely localized, while its visual pigment was similar to that of P. pyralis. These results show that both screening and visual pigments vary among species. Modeling of spectral tuning indicates that the combination of screening and visual pigments found in the retina of each species provides the best possible match of sensitivity to bioluminescent emission. This combination also produced model sensitivity spectra that closely resemble sensitivities measured either with electroretinographic or intracellular techniques. Vision in both species of Photinus appears to be evolutionarily tuned for maximum discrimination of conspecific signals from spectrally broader backgrounds. Ph. versicolor, on the other hand, appears to have a visual system that offers a compromise between maximum sensitivity to, and maximum discrimination of, their signals. Accepted: 29 September 1999     

Properties of pupil mechanisms in owl-fly Ascalaphus macaronius (Neuroptera)

Regulation of light flux by pupil mechanisms in the UV-sensitive superposition eye of owl-fly Ascalaphus macaronius (Neuroptera) was studied with a fast reflection microspectrophotometric technique. The spectral sensitivity of pupil reaction, which was calculated on the basis of changes of transient amplitude reflection, was almost identical with the one of Deilephila eye. This indicates that in spite of different life styles and spectral sensitivities of photoreceptors, pupil closing is triggered by the same photosensitive structure in both eyes. By measuring the spectra of reflected light from the Ascalaphus eye between 400 and 700 nm after different dark periods following light stimulation, it was established that the restoration of reflection was much faster in the red than in the blue spectral range. Based on this, we propose that two different pupil mechanisms with different spectral absorption characteristics are involved in light-flux regulation. Fast-reacting pupil is probably represented by screening pigment migration in the secondary pigment cells and a slow blue-absorbing system by the activity in primary pigment cells. The importance of two different pupils for the photoregeneration of visual pigment is discussed. Accepted: 1 October 1998     

Gaussian band analysis of absorption,fluorescence and photobleaching difference spectra of D1/D2/cytb-559 complex

A study of the absorption and fluorescence characteristics of the D1/D2/cytb-559 reaction centre complex of Photosystem II has been carried out by gaussian decomposition of absorption spectra both at room temperature and 72 K and of the room temperature fluorescence spectrum. A five component fit was found in which the absorption and fluorescence sub-bands could be connected by the Stepanov relation. The photobleaching and light-activated degradation in the dark of long wavelength pigments permitted a further characterisation of the absorption bands. The absorption (fluorescence) maxima of the five bands at room temperature are 660 nm (670 nm), 669 nm (675 nm), 675 nm (681 nm), 680 nm (683 nm), 681 nm (689 nm). A novel feature of this analysis is the presence of two approximately isoenergetic absorption bands near 680 nm at room temperature. The narrower one (FWHM=12.5 nm) is attributed to pheophytin while the broader band (FWHM=23 nm) is thought to be P680. The P680 band width is discussed in terms of homogeneous and site inhomogeous band broadening. The P680 fluorescence has a large Stokes shift (9 nm) and most fluorescence in the 690–700 nm range is associated with this chromophore.The three accessory pigment bands are broad (FWHM=17–24 nm) and the 660 nm gaussian is largely temperature insensitive thus indicating significant site inhomogeneous broadening.The very slight narrowing of the D1/D2/cytb-559 Q_y absorption at crytogenic temperatures is discussed in terms of the coarse spectral inhomogeneity associated with the spectral forms and the apparently large site inhomogeneous broadening of short wavelength accessory pigments.     

The effect of eye colour pigments on the action spectrum of Drosophila

In vivo absorption spectra for Drosophila melanogaster eye colour pigment classes (drosopterins and ommatins) were constructed by subtracting the whole eye electroretinographic (ERG) spectral sensitivities of cn and bw respectively from the sensitivities of white-eyed strains. In situ microspectrophotometric (MSP) absorption spectra were also obtained. Both the ERG and MSP drosopterin spectra show a visible peak at 500 nm compared to the 480 nm peak of in vitro drosopterins. For the ommatins, the ERG absorption spectrum peaks at 450 nm while the MSP spectrum peaks at 400 and 525 nm. The ERG spectrum is similar to the in vitro absorption spectrum of xanthommatin while the MSP spectrum is similar to the in vitro absorption spectrum of reduced xanthommatin. The ERG absorption spectra for the drosopterins and the ommatins yield an accurate prediction of the effect of the combined pigments in wild-type eyes. Newly emerged and 7 day post-emergence bw flies show quantitatively similar pigment absorption effects while the drosopterins depress the sensitivity of newly emerged cn flies to a greater extent than that of cn flies 7 days after emergence.     

Evolution and mechanism of spectral tuning of blue-absorbing visual pigments in butterflies

The eyes of flower-visiting butterflies are often spectrally highly complex with multiple opsin genes generated by gene duplication, providing an interesting system for a comparative study of color vision. The Small White butterfly, Pieris rapae, has duplicated blue opsins, PrB and PrV, which are expressed in the blue (λ _max = 453 nm) and violet receptors (λ _max = 425 nm), respectively. To reveal accurate absorption profiles and the molecular basis of the spectral tuning of these visual pigments, we successfully modified our honeybee opsin expression system based on HEK293s cells, and expressed PrB and PrV, the first lepidopteran opsins ever expressed in cultured cells. We reconstituted the expressed visual pigments in vitro, and analysed them spectroscopically. Both reconstituted visual pigments had two photointerconvertible states, rhodopsin and metarhodopsin, with absorption peak wavelengths 450 nm and 485 nm for PrB and 420 nm and 482 nm for PrV. We furthermore introduced site-directed mutations to the opsins and found that two amino acid substitutions, at positions 116 and 177, were crucial for the spectral tuning. This tuning mechanism appears to be specific for invertebrates and is partially shared by other pierid and lycaenid butterfly species.     

Spectral and kinetic parameters of phosphorescence of triplet chlorophyll a in the photosynthetic apparatus of plants

Spectral and kinetic parameters and quantum yield of IR phosphorescence accompanying radiative deactivation of the chlorophyll a (Chl a) triplet state were compared in pigment solutions, greening and mature plant leaves, isolated chloroplasts, and thalluses of macrophytic marine algae. On the early stages of greening just after the Shibata shift, phosphorescence is determined by the bulk Chl a molecules. According to phosphorescence measurement, the quantum yield of triplet state formation is not less than 25%. Further greening leads to a strong decrease in the phosphorescence yield. In mature leaves developing under normal irradiation conditions, the phosphorescence yield declined 1000-fold. This parameter is stable in leaves of different plant species. Three spectral forms of phosphorescence-emitting chlorophyll were revealed in the mature photosynthetic apparatus with the main emission maxima at 955, 975, and 995 nm and lifetimes ～1.9, ～1.5, and 1.1–1.3 ms. In the excitation spectra of chlorophyll phosphorescence measured in thalluses of macrophytic green and red algae, the absorption bands of Chl a and accessory pigments — carotenoids, Chl b, and phycobilins — were observed. These data suggest that phosphorescence is emitted by triplet chlorophyll molecules that are not quenched by carotenoids and correspond to short wavelength forms of Chl a coupled to the normal light harvesting pigment complex. The concentration of the phosphorescence-emitting chlorophyll molecules in chloroplasts and the contribution of these molecules to chlorophyll fluorescence were estimated. Spectral and kinetic parameters of the phosphorescence corresponding to the long wavelength fluorescence band at 737 nm were evaluated. The data indicate that phosphorescence provides unique information on the photophysics of pigment molecules, molecular organization of the photosynthetic apparatus, and mechanisms and efficiency of photodynamic stress in plants.     

The Spectral Sensitivities of Single Receptor Cells in the Lateral, Median, and Ventral Eyes of Normal and White-Eyed Limulus

Spectral sensitivity curves can be distorted by screening pigments. We have determined whether this is true for Limulus polyphemus by determining, from receptor potentials recorded using intracellular microelectrodes, spectral sensitivity curves for normal animals and for white-eyed animals (which lack screening pigment). Our results show: (a) In median ocelli, the curve for UV-sensitive receptor cells peaks at 360 nm and does not depend on the presence of screening pigment, (b) The curve for ventral eye photoreceptors is identical to that for retinular cells from the lateral eyes of white-eyed animals and peaks at 520–525 nm. (c) In normal lateral eyes, when the stimulating light passes through screening pigment, the curve indicates relatively more sensitivity in the red region of the spectrum than does the curve for white-eyed animals. Therefore, the screening pigment is probably red-transmitting, (d) In median ocelli, the curve for visible-sensitive cells peaks at 525 nm and is approximately the same whether the ocelli are from normal or white-eyed animals. However, the curve is significantly broader than that for ventral eyes and for lateral eyes from white-eyed animals.     

Photosynthetic action spectra of marine algae

A polarographic oxygen determination, with tissue in direct contact with a stationary platinum electrode, has been used to measure the photosynthetic response of marine algae. These were exposed to monochromatic light, of equal energy, at some 35 points through the visible spectrum (derived from a monochromator). Ulva and Monostroma (green algae) show action spectra which correspond very closely to their absorption spectra. Coilodesme (a brown alga) shows almost as good correspondence, including the spectral region absorbed by the carotenoid, fucoxanthin. In green and brown algae, light absorbed by both chlorophyll and carotenoids seems photosynthetically effective, although some inactive absorption by carotenoids is indicated. Action spectra for a wide variety of red algae, however, show marked deviations from their corresponding absorption spectra. The photosynthetic rates are high in the spectral regions absorbed by the water-soluble "phycobilin" pigments (phycoerythrin and phycocyanin), while the light absorbed by chlorophyll and carotenoids is poorly utilized for oxygen production. In red algae containing chiefly phycoerythrin, the action spectrum closely resembles that of the water-extracted pigment, with peaks corresponding to its absorption maxima (495, 540, and 565 mµ). Such algae include Delesseria, Schizymenia, and Porphyrella. In the genus Porphyra, there is a series P. nereocystis, P. naiadum, and P. perforata, with increasingly more phycocyanin and less phycoerythrin: the action spectra reflect this, with increasing activity in the orange-red region (600 to 640 mµ) where phycocyanin absorbs. In all these red algae, photosynthesis is almost minimal at 435 mµ and 675 mµ, where chlorophyll shows maximum absorption. Although the chlorophylls (and carotenoids) are present in quantities comparable to the green algae, their function is apparently not that of a primary light absorber; this role is taken over by the phycobilins. In this respect the red algae (Rhodophyta) appear unique among photosynthetic plants.     

Profiles of photosynthetic pigment accumulation and expression of photosynthesis-related genes in the marine cyanobacteria Synechococcus sp.: Effects of LED wavelengths

Light quality is a significant environmental factor that influences photosynthetic pigments in cyanobacteria. In the present study, we illuminated the marine cyanobacteria Synechococcus sp. with white (350 ～ 700 nm), red (630 nm), green (530 nm), and blue (450 nm) light emitting diodes (LEDs) and measured pigment levels (chlorophyll, carotenoid, and phycobiliprotein) and expression of photosynthesis-related genes (pebA, psbB, and psaE). The amount of photosynthetic pigments (total pigments, chlorophyll, and phycobiliproteins) was higher in the green and blue LED groups than in the white and red LED groups after 8 days of culture. The cells were prepared in a 1.5 mL solution for the analysis of the total pigments, chlorophyll, and carotenoid, and in a 2 mL for analysis of phycobiliproteins. The mRNA expression levels of pebA and psbB significantly increased after 8 days of cultivation under green and blue light, while the mRNA expression levels of psaE decreased. These results indicate that green and blue light increase the accumulation of photosynthetic pigments. In contrast red light induced mRNA expression of psaE and stimulated cell growth in Synechococcus sp.     

pH值及金属离子对樟树果实红色素吸收光谱的影响

以樟树果实为材料,运用800～400 nm光谱扫描分析了pH值、金属离子、加热时间及温度、光照等稳定性因子对樟树果实红色素吸收光谱的影响。结果表明:最大吸收波长是517 nm,酸性条件对色素吸收光谱无明显影响,当pH为8.6的强碱性环境,红色素结构变化,最大吸收峰漂移至402 nm;100℃内,色素性质稳定,当加热时间延长至120 min,最大吸收峰在512.5 nm;红色素耐光性较好,但避光更利于保存;金属离子对红色素吸收光谱的影响强弱为:Fe3+>Al3+>Mg2+>Ca2+>Na+>K+,Fe3+短时内引起色素变色、发生沉淀,最大吸收峰漂移,浓度越高影响越大,Na+、K+对红色素吸收光谱无明显影响。     

Comparative analysis of field and laboratory spectral reflectances of benthic diatoms with a modified Gaussian model approach

Benthic diatom biomass on an intertidal mudflat was estimated by field spectrometry, a non-intrusive optical method operating in the visible-infrared wavelength range. Spectral reflectance (400-900 nm) of natural assemblages was related to the amount of principal photosynthetic and accessory pigments measured by high-performance liquid chromatography (HPLC) in the top 2 mm of sediment. Relationships established in situ were compared with those obtained in the laboratory with monospecific cultures. However, to compare both types of reflectance spectra differing in their overall shape (continuum) and to isolate the pigment absorption features from other effects, we applied hyperspectral data processing, the Modified Gaussian Model (MGM), to remove the continuum and model the main absorption bands by a succession of Gaussian curves. The Gaussian band depths associated with the absorption by chlorophyll a and the pigments specific to diatoms (chlorophyll c, fucoxanthine, diadinoxanthin) displayed linear relationships with the logarithm of chlorophyll a.The most relevant spectral feature to quantify the microalgal biomass in situ was the 632 nm absorption band associated with chlorophyll c. A significant but equivocal statistical relationship was obtained at 675 nm, due to the overlapping absorption by chlorophyll breakdown products (pheophytine a and pheophorbide a) present in all the samples analyzed in the field. Fucoxanthin absorption at 550 nm can become an indicator of chlorophyll a biomass for benthic diatoms, but the effect of seasonal photophysiological adaptation should be considered. The comparative analysis of field and laboratory reflectance showed that the chlorophyll a present in the top 2 mm was not a good estimator of the photosynthetic active biomass (PAB) and that a shallower depth should be sampled in muddy sediments colonized by microphytobenthos. The approach of MGM band depth retrieval offers the possibility of quantifying PAB and has the potential to characterize (by accessory pigments) microalgae present in the photic zone of different sediment types, removing albedo variability due to grain-size scattering effects or sediment moisture content. Its accuracy relies, however, on an appropriate ground-truth sampling, excluding the contribution of redundant pigments not detected by the sensor.