# 2.3: Light spectroscopy - Biology

Spectrophotometers measure the amount of light absorbed by a sample at a particular wavelength. Measurements are usually made at a wavelength that is close to the absorbance maximum for the molecule of interest in the sample.

The diagram below shows the elements present in a typical spectrophotometer. The light sources used in most spectrophotometers emit either ultraviolet or visible light. Light
(Io) passes from a source to a monochromator, which can be adjusted to allow only light of a defined wavelength to pass through. The monochromatic (I) light then passes through a cuvette containing the sample to a detector.

The spectrophotometer compares the fraction of light passing through the monochromator (I0) to the light reaching the detector (I) and computes the transmittance (T) as I/I0. Absorbance (A) is a logarithmic function of the transmittance and is calculated as:

A = log10(1/T) = log10(I0/I)

Spectrophotometers can express data as either % transmittance or absorbance. Most investigators prefer to collect absorbance values, because the absorbance of a compound is directly proportional to its concentration. Recall the Lambert-Beer Law, traditionally expressed as:

A =(varepsilon)b C

where (varepsilon) is the molar extinction coefficient of a compound, b is the length of the light path through the sample, and C is the molar concentration of the compound. Cuvettes are formulated to have a 1 cm light path, and the molar extinction coefficient is expressed as L/moles-cm. Consequently, absorbance is a unitless value.

## Spectra and What They Can Tell Us

A spectrum is simply a chart or a graph that shows the intensity of light being emitted over a range of energies. Have you ever seen a spectrum before? Probably. Nature makes beautiful ones we call rainbows. Sunlight sent through raindrops is spread out to display its various colors (the different colors are just the way our eyes perceive radiation with slightly different energies).

Spectroscopy can be very useful in helping scientists understand how an object like a black hole, neutron star, or active galaxy produces light, how fast it is moving, and what elements it is composed of. Spectra can be produced for any energy of light, from low-energy radio waves to very high-energy gamma rays.

Each spectrum holds a wide variety of information. For instance, there are many different mechanisms by which an object, like a star, can produce light. Each of these mechanisms has a characteristic spectrum.

## Implementation of a six-around-one optical probe based on diffuse light spectroscopy for study of cerebral properties in a murine mouse model of autism spectrum disorder

Light reflectance spectroscopy (LRS) is a multispectral technique, sensitive to the absorption and scattering properties of biological molecules in tissues. It is used as a noninvasive tool to extract quantitative physiological information from human tissues and organs. A near-infrared LRS based on a single optical probe was used to monitor changes in optical and hemodynamic parameters in a mouse model of autism. A murine model of autism induced by developmental exposure to valproic acid (VPA) was used. Since autism could be attributed to neuroanatomical changes, we hypothesize that these changes can be detected using the LRS because spectral properties depend on both molecular composition and structural changes. The fiber-optic probe in the setup consisted of seven small optical fibers: six fibers for illumination placed in a circular manner around a central single collection fiber. Overall, measurements demonstrate changes in diffused reflectance spectra, cerebral optical tissue properties (absorption and scattering), and chromophore levels. Furthermore, we were able to identify differences between male and female groups. Finally, the effectiveness of S-Adenosylmethionine as a drug therapy was studied and found to improve the hemodynamic outcome. For the first time, to the best of our knowledge, the LRS is utilized to study variations in brain parameters in the VPA autism model mice through an intact scalp.

© 2020 Optical Society of America

Huiyi Cheng, Jie Yu, Lingyu Xu, and Jun Li
Biomed. Opt. Express 10(3) 1383-1392 (2019)

Huilin Zhu, Yuebo Fan, Huan Guo, Dan Huang, and Sailing He
Biomed. Opt. Express 5(4) 1262-1274 (2014)

Oren Shaul, Michal Fanrazi-Kahana, Omri Meitav, Gad A. Pinhasi, and David Abookasis
Appl. Opt. 56(32) 8880-8886 (2017)

Jun Li, Lina Qiu, Lingyu Xu, Ernest V. Pedapati, Craig A. Erickson, and Ulas Sunar
Biomed. Opt. Express 7(10) 3871-3881 (2016)

Huilin Zhu, Jun Li, Yuebo Fan, Xinge Li, Dan Huang, and Sailing He
Biomed. Opt. Express 6(3) 690-701 (2015)

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## STARK SPECTROSCOPY: Applications in Chemistry, Biology, and Materials Science

AbstractStark spectroscopy has been applied to a wide range of molecular systems and materials. A generally useful method for obtaining electronic and vibrational Stark spectra that does not require sophisticated equipment is described. By working with frozen glasses it is possible to study nearly any molecular system, including ions and proteins. Quantitative analysis of the spectra provides information on the change in dipole moment and polarizability associated with a transition. The change in dipole moment reflects the degree of charge separation for a transition, a quantity of interest to a variety of fields. The polarizability change describes the sensitivity of a transition to an electrostatic field such as that found in a protein or an ordered synthetic material. Applications to donor-acceptor polyenes, transition metal complexes (metal-to-ligand and metal-to-metal mixed valence transitions), and nonphotosynthetic biological systems are reviewed.

## Discussion

Our results suggest that the installation of broader spectrum lighting technologies in artificially lit habitats is likely to improve the ability of animals to detect light reflected from objects in their environment at night, and has the potential to generate greater disparities in this ability between different classes of animal. These improvements in object detection under broad spectrum street lights are likely to affect the execution of visually guided behaviours in animals, altering their normal activity times and spatially extending or fragmenting habitats. All three broad spectrum lighting technologies provided significant improvements in the % λ0.5 range in comparison to narrow spectrum LPS lamps. MH lamps provided the greatest improvements in all five taxonomic classes. Hence, where these are in use, a greater variety of objects reflecting light in different regions of the light spectrum will appear brighter and more colourful to animals compared with alternative street lamp technologies. While LPS lamps illuminate objects reflecting light across the smallest region of the light spectrum, our results suggest that in areas illuminated by LPS lamps, birds and mammals are better able to detect objects that reflect light in this region compared to arachnids, insects and reptiles. The introduction of broader spectrum technologies, however, increases the number, and the magnitude of the differences between animal classes, in the proportion of the visually detectable light spectrum illuminated, with mammals and birds displaying the largest improvements. Most mammals possess dichromatic vision spanning a less extended range of the light spectrum in comparison to birds, reptiles, arachnids and insects (Fig. 2a see Table S1) that typically can detect light at wavelengths below 400 nm (UV) (Tovée, 1995 Briscoe & Chittka, 2001 Hart & Hunt, 2007 Osorio & Vorobyev, 2008 ). Birds do possess UV sensitive photoreceptors, but their sensitivity extends less into the shorter wavelengths compared to insects, arachnids and reptiles (Fig. 2a). Broad spectrum lamp types therefore stimulate a larger percentage of the λ0.5 range in mammals and birds in general, compared with other classes of animal, improving their ability to perform visually guided behaviours with greater acuity and potentially upsetting the balance of interspecific interactions.

Our results provide an overview of how shifting artificial light spectra are likely to affect visually guided behaviours in broad taxonomic groups of animal. However, the λ0.5 range of individual species can be variable within each taxonomic group, and therefore caution should be exercised when applying the results of a group in general to any one specific species within that group. For example, the number of photoreceptor types in insect eyes is variable between different orders (Table S1) giving rise to variation in the proportion of λ0.5 range illuminated by each type of artificial light. In addition, the number of species for which λmax values are available in the literature varies between taxonomic groups (Table S1), and while the main results of this study are unlikely to be affected, the λ0.5 range will inevitably adjust as data become available for more species and additional photoreceptors in those groups which are not currently well investigated (for example the arachnids). These results are not therefore conclusive, rather they should be considered as a platform of predictions which incentivises further studies into the impact of broadening artificial light spectra on visually guided behaviours in animals.

The ecological impacts of artificially lighting the nocturnal environment are increasingly being recognized (Frank, 2006 Stone et al., 2012 Titulaer et al., 2012 ), with some studies drawing attention to the potential impact of shifting spectral signatures (Eisenbeis, 2006 Stone et al., 2012 ). This study has highlighted that such changes may be affecting visually guided behaviours in species across the animal kingdom. The range of potential impacts are diverse and may include extending the times of foraging and sexual competition of diurnal and crepuscular animals into the night (Robertson & Monteiro, 2005 Somanathan et al., 2009 Titulaer et al., 2012 ), improving both prey detection and predator avoidance (Roth & Kelber, 2004 ), changing the ability of organisms to navigate around their environment (Warrant et al., 2004 , Somanathan et al., 2008 Stone et al., 2009 van Langevelde et al., 2011 ) and affecting the ability of pollinating species to detect nectar resources (Kelber et al., 2002 Hempel de Ibarra & Vorobyev, 2009 ). Whether broadening artificial light spectra will elicit positive or negative species responses is likely to depend on the species and the behaviour being considered. For example, the presence of LED lighting increases feeding rates in nesting Great Tits Parus major (Titulaer et al., 2012 ), while the bat Rhinolophus hipposideros avoids areas lit by HPS and LED lighting (Stone et al., 2009 , 2012 ) potentially due to perceived predation risk (Rydell, 1992 ). Metal Halide (MH) lamps are likely to provide the largest improvements in animal vision because they emit light that is both broad and contains UV in its spectral composition. Many of the above tasks depend on the perception of UV light reflected from objects by animals that can detect light at these wavelengths. Hence, the introduction of broader spectrum lighting technologies containing UV may have more profound consequences for biological systems than non-UV broad spectrum lighting technologies. All three broad spectrum technologies, however, create larger disparities in % λ0.5 between animal groups compared with narrow spectrum LPS lamps, and so have greater potential to alter the balance of interspecific interactions in the environment. Evaluating the direct environmental impacts of each of these different lamp types is clearly essential in a world where the artificially lit night-time environment is increasingly becoming ‘white’.