Photosynthetica X:X | DOI: 10.32615/ps.2026.033

Determination of chlorophyll and carotenoid content in Mediterranean riparian forest species using visible spectroscopy

M. MARTÍNEZ MENA1, M.D.L. SOTO ROSALES2, D. GARCÍA DE JALÓN1
1 Hydrobiology Group, Department of Systems and Natural Resources, Higher Technical School of Forestry, Forest and Natural Environment, Polytechnic University of Madrid, University City, 28040 Madrid, Spain
2 National Polytechnic Institute, Higher School of Economics, Plan de Agua Prieta 66, Plutarco Elías Calles, Miguel Hidalgo, 11350 Mexico City, Mexico

Leaf-level optical measurements are essential for monitoring Mediterranean riparian forests. This study used visible spectroscopy (400-750 nm) to determine and quantify leaf pigments in five species: Salix salviifolia, Fraxinus angustifolia, Alnus glutinosa, Populus nigra, and Prunus avium. We followed Hansmann's method for sample processing and quantified chlorophyll using Arnon's equation. We collected samples from sun-exposed leaves in spring under standardized conditions. Results indicated that Alnus glutinosa and Populus nigra exhibited the highest chlorophyll a concentrations [1.35 and 1.29 mg g-1 (FM)], followed by Fraxinus angustifolia [1.17 mg g-1(FM)], Salix salviifolia [0.61 mg g-1(FM)], and Prunus avium [0.25 mg g-1(FM)]. The chlorophyll a/b ratio exceeded 2.0 in most species, peaking in Salix salviifolia (2.90). These findings reveal significant functional differences in pigment composition, confirming the utility of this method for monitoring the physiological status of the vegetation in the Manzanares River, Spain.

Additional key words: chlorophyll a; chlorophyll a, b ratio; Manzanares Rivercstyle

Received: March 20, 2026; Revised: August 17, 2026; Accepted: September 21, 2026; Prepublished online: October 13, 2026 

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    References

    1. Abdullah H., Darvishzadeh R., Skidmore A.K. et al.: European spruce bark beetle (Ips typographus, L.) green attack affects foliar reflectance and biochemical properties. - Int. J. Appl. Earth Obs. Geoinf. 64: 199-209, 2018. Go to original source...
    2. AEMET: Spanish State Meteorological Agency: forecasts, advisories, observations, situation report, 2024. Available at: https://www.aemet.es/es/datos_abiertos. [In Spanish]
    3. Alaboodi A.S., Kadhim S.A., Hussein A.S.: Ultraviolet-visible spectroscopy, importance, principle, structure and most important applications: a study review. - Int. J. Nov. Res. Phys. Chem. Math. 12: 53-60, 2025.
    4. Arnon D.I.: Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. - Plant Physiol. 24: 1-15, 1949. Go to original source...
    5. Barbosa T.K.M., Fiorio P.R., Calaboni C. et al.: Lisianthus (Eustoma grandiflorum) leaf degradation analysis in the postharvest by VIS-NIR-SWIR reflectance spectroscopy. - Ciênc. Rural 54: e20230143, 2024. Go to original source...
    6. Bednaøíková M., Gauslaa Y., Solhaug K.A.: Non-invasive monitoring of photosynthetic activity and water content in forest lichens using spectral reflectance data and RGB colors from photographs. - Fungal Ecol. 62: 101224, 2023. Go to original source...
    7. Camarero J.J., Colangelo M., Rodríguez-González P.M.: Tree growth, wood anatomy and carbon and oxygen isotopes responses to drought in Mediterranean riparian forests. - Forest Ecol. Manag. 529: 120710, 2023. Go to original source...
    8. Camarero J.J., Gazol A., Fernández-Cortés Á., Colangelo M.: Riparian field elm (Ulmus minor) is sensitive to drought, and fast-growing trees are susceptible to Dutch elm disease. - Forest Ecol. Manag. 594: 122948, 2025. Go to original source...
    9. Casotti C.G., Kiffer Jr W.P., Costa L.C. et al.: Assessing the importance of riparian zones conservation for leaf decomposition in streams. - Natur. Conserv. 13: 178-182, 2015. Go to original source...
    10. Castroviejo S., Aedo C., Cirujano S. et al.: [Flora Ibérica: Vascular Plants of the Iberian Peninsula and the Balearic Islands.] Pp. 784. CSIC, Madrid 2020. [In Spanish]
    11. Chen M., Blankenship R.E.: Expanding the solar spectrum used by photosynthesis. - Trends Plant Sci. 16: 427-431, 2011. Go to original source...
    12. Chen M., Schliep M., Willows R.D. et al.: A red-shifted chlorophyll. - Science 329: 1318-1319, 2010. Go to original source...
    13. Cid Rusiñol M., Sánchez Fayos Calabuig T., Villacañas Beades S.: [Special plan for the protection and improvement of the green infrastructure of the metropolitan forest of Madrid. Lot 1. General Directorate of Strategic Planning.] Ayuntamiento de Madrid, Madrid 2023. Available at: https://servpub.madrid.es/VECSV_WBCONSULTA/VerificarCove.do. [In Spanish]
    14. Croft H., Chen J.M., Luo X. et al.: Leaf chlorophyll content as a proxy for leaf photosynthetic capacity. - Glob. Change Biol. 23: 3513-3524, 2017. Go to original source...
    15. Del Tánago M.G., Martínez-Fernández V., Aguiar F.C. et al.: Improving river hydromorphological assessment through better integration of riparian vegetation: Scientific evidence and guidelines. - J. Environ. Manage. 292: 112730, 2021. Go to original source...
    16. Deng X., Greet J., Jones C.S.: Soil moisture influences the root characteristics of a herbaceous riparian plant along a regulated river. - Plant Ecol. 225: 25-36, 2024. Go to original source...
    17. Díaz-Alba D., Henry A.L., de Jalón D.G. et al.: Salix regeneration in fluvial landscapes: Empirical findings based on a systematic review. - Ecol. Eng. 193: 107010, 2023. Go to original source...
    18. Díaz-Redondo M., Marchamalo M., Morcillo F., King J.J.: Naturalising a heavily modified urban river: Initial habitat evolution in the Manzanares River (Madrid, Spain). - River Res. Appl. 38: 727-737, 2022. Go to original source...
    19. Du S., Zhao D., Guan L. et al.: PROSPECT-DP: A coupled leaf optical properties model for improving leaf spectra simulation in the red-edge domain by excluding the ChlF effect. - Int. J. Appl. Earth Obs. Geoinf. 141: 104651, 2025. Go to original source...
    20. Gao R.-Y., Zou J.-W., Shi Y.-P. et al.: The Q-band energetics and relaxation of chlorophylls a and b as revealed by visible-to-near infrared time-resolved absorption spectroscopy. - J. Phys. Chem. Lett. 16: 789-794, 2025. Go to original source...
    21. Gong L., Zhu C., Luo Y., Fu X.: Spectral reflectance reconstruction from red-green-blue (RGB) images for chlorophyll content detection. - Appl. Spectrosc. 77: 200-209, 2023. Go to original source...
    22. González E., Martínez-Fernández V., Shafroth P.B. et al.: Regeneration of Salicaceae riparian forests in the Northern Hemisphere: a new framework and management tool. - J. Environ. Manage. 218: 374-387, 2018. Go to original source...
    23. Gran K.B., Tal M., Wartman E.D.: Co-evolution of riparian vegetation and channel dynamics in an aggrading braided river system, Mount Pinatubo, Philippines. - Earth Surf. Proc. Land. 40: 1101-1115, 2015. Go to original source...
    24. Gurnell A.M., Scott S.J., England J. et al.: Assessing river condition: A multiscale approach designed for operational application in the context of biodiversity net gain. - River Res. Appl. 36: 1559-1578, 2020. Go to original source...
    25. Havrdová A., Douda J., Doudová J.: Threats, biodiversity drivers and restoration in temperate floodplain forests related to spatial scales. - Sci. Total Environ. 854: 158743, 2023. Go to original source...
    26. Houborg R., Fisher J.B., Skidmore A.K.: Advances in remote sensing of vegetation function and traits. - J. Appl. Earth Obs. Geoinf. 43: 1-6, 2015. Go to original source...
    27. IUSS Working Group WRB: World Reference Base for Soil Resources 2014. International Soil Classification System for Naming Soils and Creating Legends for Soil Maps. World Soil Resources Reports No. 106. Pp. 203. FAO, Rome 2015. Available at: https://openknowledge.fao.org/handle/20.500.14283/i3794en.
    28. Kior A., Yudina L., Zolin Y. et al.: RGB imaging as a tool for remote sensing of characteristics of terrestrial plants: a review. - Plants-Basel 13: 1262, 2024. Go to original source...
    29. Lichtenthaler H.K., Babani F.: Contents of photosynthetic pigments and ratios of chlorophyll a/b and chlorophylls to carotenoids (a+b)/(x+c) in C4 plants as compared to C3 plants. - Photosynthetica 60: 3-9, 2022. Go to original source...
    30. López González G.A.: [The Trees and Shrubs of the Iberian Peninsula and the Balearic Islands.] 2nd Edition. 2 Volumes. Pp. 1731. Ediciones Mundi-Prensa, Madrid 2006. [In Spanish]
    31. Lozanovska I., Bejarano M.D., Martins M.J. et al.: The functional diversity of riparian woody vegetation is less affected by river regulation in the Mediterranean than in the boreal region. - Front. Plant Sci. 11: 857, 2020. Go to original source...
    32. Mackinney G.: Absorption of light by chlorophyll solutions. - J. Biol. Chem. 140: 315-322, 1941. Go to original source...
    33. Martínez-Fernández V., Van Oorschot M., De Smit J. et al.: Modelling feedbacks between geomorphological and riparian vegetation responses under climate change in a Mediterranean context. - Earth Surf. Proc. Land. 43: 1825-1835, 2018. Go to original source...
    34. Milne B.F., Toker Y., Rubio A., Nielsen S.B.: Unraveling the intrinsic color of chlorophyll. - Angew. Chem. Int. Ed. 54: 2170-2173, 2015. Go to original source...
    35. Montes C., Llorca A., Sterling A.: [Guidelines for the Ecological Recovery of the Middle Section of the Manzanares River.] 1st Edition. Pp. 139. Canal de Isabel II, Madrid 1987. [In Spanish]
    36. Mulero G., Bacher H., Kleiner U. et al.: Spectral estimation of in vivo wheat chlorophyll a/b ratio under contrasting water availabilities. - Remote Sens. 14: 2585, 2022. Go to original source...
    37. Newberry R.W., Raines R.T.: The n→ π* interaction. - Acc. Chem. Res. 50: 1838-1846, 2017. Go to original source...
    38. Pérez-Patricio M., Camas-Anzueto J.L., Sanchez-Alegría A. et al.: Optical method for estimating the chlorophyll contents in plant leaves. - Sensors 18: 650, 2018. Go to original source...
    39. Qiu N.W., Jiang D.C., Wang X.S. et al.: Advances in the members and biosynthesis of chlorophyll family. - Photosynthetica 57: 974-984, 2019. Go to original source...
    40. Regodón J.P.: [Geological, Hydrogeological, and Mining Guide of the Province of Madrid.] 1st Edition. Memorias del Instituto Geológico y Minero de España, Madrid 1970. [In Spanish]
    41. Riis T., Kelly-Quinn M., Aguiar F.C. et al.: Global overview of ecosystem services provided by riparian vegetation. - BioScience 70: 501-514, 2020. Go to original source...
    42. Ríos Cortés G.R.C., Ramírez Aguilar D., Bobadilla Reyes C.Q. et al.: Removal of chlorophylls present in stevia (Stevia rebaudiana Bertoni) extracts by activated carbon adsorption and precipitation with lime food grade. - Acta Univ. 27: 67-75, 2017. Go to original source...
    43. Rodríguez-González P.M., Stella J.C., Campelo F. et al.: Subsidy or stress? Tree structure and growth in wetland forests along a hydrological gradient in Southern Europe. - Forest Ecol. Manag. 259: 2015-2025, 2010. Go to original source...
    44. Sabo J.L., Sponseller R., Dixon M. et al.: Riparian areas increase regional species richness by harboring different species, not more species. - Ecology 86: 56-62, 2005. Go to original source...
    45. Santiago J.M., García de Jalón D.: Heat recovery as a tool for reducing the thermal impact of effluents from wastewater treatment plants. - Sustainability 18: 3879, 2026. Go to original source...
    46. Sinclair T., Craig P., Maltby L.L.: Climate warming shifts riverine macroinvertebrate communities to be more sensitive to chemical pollutants. - Glob. Change Biol. 30: e17254, 2024. Go to original source...
    47. Stella J.C., Bendix J.: Multiple stressors in riparian ecosystems. -In: Sabater S., Elosegi A., Ludwig R. (Eds.): Multiple Stressors in River Ecosystems: Status, Impacts and Prospects for the Future. Pp. 81-110. Elsevier, San Diego 2019. Go to original source...
    48. Taniguchi M., Lindsey J.S.: Absorption and fluorescence spectral database of chlorophylls and analogues. - Photochem. Photobiol. 97: 136-165, 2021. Go to original source...
    49. Yan Z., Li P., Li Z. et al.: Dynamic response of soil enzyme activities in riparian zones: Insights from vegetation cover. - J. Soil Sci. Plant Nutr. 25: 1597-1611, 2025. Go to original source...
    50. Zahir S.A.D.M., Omar A.F., Jamlos M.F. et al.: A review of visible and near-infrared (Vis-NIR) spectroscopy application in plant stress detection. - Sensor. Actuat. A Phys. 338: 113468, 2022. Go to original source...
    51. Zahn C., Nag P., Stensitzki T. et al.: Orientation of the electronic Qy transition dipole moment in chlorophyll a. - J. Phys. Chem. B 130: 5488-5494, 2026. Go to original source...

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