BACKGROUND: Experimental determination of the extent and rate of transport of liquid humates supplied to plants is critical in testing physiological effects of such biostimulants which are often supplied as foliar sprays. Therefore, an original experimental method for the qualitative investigation and quantitative description of the penetration of humates through plant cuticles is proposed, tested, and evaluated. RESULTS: The proposed method involves the isolation of model plant leaf cuticles and the subsequent in vitro evaluation of cuticular humate transport. The employed novel methodology is based on a simple diffusion couple arrangement involving continuous spectrophotometric determination of the amount of penetrated humate in a hydrogel diffusion medium. Prunus laurocerasus leaf cuticles were isolated by chemical and enzymatic treatment and the rate of cuticular penetration of a commercial humate (lignohumate) was estimated over time in quantitative and qualitative terms. Different rates of lignohumate transport were determined for abaxial and adaxial leaf cuticles also in relation to the different cuticular extraction methods tested. CONCLUSIONS: The proposed methodology represents a simple and cheap experimental tool for the study on the trans-cuticular penetration of humic-based biostimulants.

Institute of Physical and Applied Chemistry Brno University of Technology Faculty of Chemistry Purkynova 464 118 612 00 Brno Czech Republic

Materials Research Centre Brno University of Technology Faculty of Chemistry Purkynova 464 118 612 00 Brno Czech Republic

See more in PubMed

Neshev N, Manolov I. Content and uptake of nutrients with plant biomass of potatoes depending on potassium fertilization. Agric Agric Sci Procedia. 2015 doi: 10.1016/j.aaspro.2015.08.039. DOI

Peigne J, Vian JF, Payet V, Saby NPA. Soil fertility after 10 years of conservation tillage in organic farming. Soil Tillage Res. 2018 doi: 10.1016/j.still.2017.09.008. DOI

Okuda A, Kawasaki T, Yamada Y. Foliar absorption of nutrients. Soil Sci Plant Nutr. 1960 doi: 10.1080/00380768.1960.10430928. DOI

Fernandez V, Eichert T. Uptake of hydrophilic solutes through plant leaves: current state of knowledge and perspectives of foliar fertilization. Crit Rev Plant Sci. 2009 doi: 10.1080/07352680902743069. DOI

Fageria NK, Barbosa Filho MP, Moreira A, Guimaraes CM. Foliar fertilization of crop plants. J Plant Nutr. 2009 doi: 10.1080/01904160902872826. DOI

Mandic V, Simic A, Krnjaja Z, Bijelic Z, Tomic A, Stanojkovic D, Ruzic Music D. Effect of foliar fertilization on soybean grain yield. Biotechnol Anim Husb. 2015 doi: 10.2298/BAH1501133M. DOI

Li M, Wang S, Tian X, Li S, Chen Y, Jia Z, Liu K, Zhai A. Zinc and iron concentrations in grain milling fractions through combined foliar applications of Zn and macronutrients. Field Crop Res. 2016 doi: 10.1016/j.fcr.2015.12.018. DOI

Chamel A, Gambonnet B. Sorption and diffusion of an ethoxylated stearic alcohol and an ethoxylated stearic amine into and through isolated plant cuticles. Chemosphere. 1997 doi: 10.1016/S0045-6535(97)00033-7. DOI

Liu H, Shao B, Long X, Yao Y, Meng Q. Foliar penetration enhanced by biosurfactant rhamnolipid. Colloid Surf B. 2016 doi: 10.1016/j.colsurfb.2016.05.058. PubMed DOI

Solel Z, Edgington LV. Transcuticular movement of fungicides. Phytophatology. 1972 doi: 10.1094/Phyto-63-505. DOI

Chamel A, Vitton N. Sorption and diffusion of C14-atrazine through isolated plant cuticles. Chemosphere. 1996 doi: 10.1016/0045-6535(96)00241-X. DOI

Wang CJ, Liu ZQ. Foliar uptake of pesticides—present status and future challenge. Pestic Biochem Phys. 2007 doi: 10.1016/j.pestbp.2006.04.004. DOI

Zelena V, Veverka K. Effect of surfactants and liquid fertilizers on transcuticular penetration of fungicides. Plant Prot Sci. 2007 doi: 10.17221/2236-PPS. DOI

Khayet M, Fernandez V. Estimation of the solubility parameters of model plant surfaces and agrochemicals: a valuable tool for understanding plant surface interactions. Theor Biol Med Model. 2012 doi: 10.1186/1742-4682-9-45. PubMed DOI PMC

Khorram MS, Zhang Q, Lin D, Zheng Y, Fang H, Yu Y. Biochar: a review of its impact on pesticide behavior in soil environments and its potential applications. J Environ Sci. 2016 doi: 10.1016/j.jes.2015.12.027. PubMed DOI

Martin JT, Juniper BE. The cuticles of plants. New York: St. Martin’s Press; 1970.

Riederer M, Schreiber L. Protecting against water loss: analysis of the barrier properties of plant cuticles. J Exp Bot. 2001 doi: 10.1093/jexbot/52.363.2023. PubMed DOI

Barel D. Foliar application of phosphorus compounds. Doctoral thesis, Iowa State University, USA; 1975. p. 1–345.

Villena JF, Dominguez E, Heredia A. Monitoring biopolymers present in plant cuticles by FT-IR. J Plant Physiol. 1999 doi: 10.1016/S0176-1617(00)80083-8. DOI

Pollard M, Beisson F, Li Y, Ohlrogge JB. Building lipid barriers: biosynthesis of cutin and suberin. Trends Plant Sci. 2008 doi: 10.1016/j.tplants.2008.03.003. PubMed DOI

Yeats TH, Rose JK. The formation and function of plant cuticles. Plant Physiol. 2013 doi: 10.1104/pp.113.222737. PubMed DOI PMC

Riederer M, Müller C. Biology of the plant cuticle. Annual plant reviews. Oxford: Blackwell; 2006.

Gutschick VP. Biotic and abiotic consequences of differences in leaf structure. New Phytol. 1999 doi: 10.1046/j.1469-8137.1999.00423.x. DOI

Zeisler-Diehl V, Migdal B, Schreiber L. Quantitative characterization of cuticular barrier properties: methods, requirements, and problems. J Exp Bot. 2017 doi: 10.1093/jxb/erx282. PubMed DOI

Orgell WH. The isolation of plant cuticle with pectic enzymes. Plant Physiol. 1955;30:78–80. doi: 10.1104/pp.30.1.78. PubMed DOI PMC

Holloway PJ, Baker EA. Isolation of plant cuticles with zinc chloride-hydrochloric acid solution (short communication) Plant Physiol. 1968 doi: 10.1104/pp.43.11.1878. PubMed DOI PMC

Solel Z. The systematic fungicidal effect of benzimidazole derivatives and thiophanate against Cercospora leaf spot of sugar beet. Phytopathology. 1970 doi: 10.1094/Phyto-60-1186. DOI

Edgington LV, Buchenauer H, Grossmann F. Bioassay and transcuticular movement of systematic fungicides. Pestic Sci. 1973 doi: 10.1002/ps.2780040517. DOI

Schonherr J, Lendzian K. A simple and inexpensive method of measuring water permeability of isolated plant cuticular membranes. Z Pflanzenphysiol. 1981 doi: 10.1016/S0044-328X(81)80203-6. DOI

Yamada Y, Wittwer SH, Bukovac MJ. Penetration of organic compounds through isolated cuticular membranes with special reference to C14 urea. Plant Physiol. 1964 doi: 10.1104/pp.40.1.170. PubMed DOI PMC

Darlington WA, Cirulis N. Permeability of apricot leaf cuticle. Plant Physiol. 1963 doi: 10.1104/pp.38.4.462. PubMed DOI PMC

Wittwer SH, Lundahl WS. Autoradiography as an acid in determining the grass absorption and utilization of foliar applied nutrients. Plant Physiol. 1951;26:792–797. doi: 10.1104/pp.26.4.792. PubMed DOI PMC

Eggert R, Kardos LT, Smith RD. The relative absorption of phosphorus by apple trees from foliar sprays and from soil applications of fertilizer using radioactive tracers. Proc Am Soc Hortic Sci. 1952;60:75–86.

Tukey HB, Ticknor RL, Hinsvark ON, Wittwer SH. Absorption of nutrients by stems and branches of woody plants. Science. 1952 doi: 10.1126/science.116.3007.167. PubMed DOI

Bargel H, Koch K, Cerman Z, Neinhuis Ch. Structure-function relationships of the plant cuticle and cuticular waxes—a smart material? Funct Plant Biol. 2006 doi: 10.1071/FP06139. PubMed DOI

Bukovac MH, Wittwer SH. Absorption and mobility of foliar applied nutrients. Plant Physiol. 1957 doi: 10.1104/pp.32.5.428. PubMed DOI PMC

Yamada Y, Wittwer SH, Bukovac MJ. Penetration of ions through isolated cuticles. Plant Physiol. 1963 doi: 10.1104/pp.39.1.28. PubMed DOI PMC

Yu JH, Lim HK, Choi GJ, Cho KY, Kim JH. A new method for assessing foliar uptake of fungicides using Congo Red as a tracer. Pest Manag Sci. 2001 doi: 10.1002/ps.327. PubMed DOI

Pinton R, Cesco S, Santi S, Varanini Z. Soil humic substances stimulate proton release by intact oat seedling roots. J Plant Nutr. 1997 doi: 10.1080/01904169709365301. DOI

Chen Y, Clapp CE, Magen H, Cline VW. Stimulation of plant growth by humic substances: effects on iron availability. In: Ghabbour EA, Davies G, editors. Understanding humic substances: advanced methods, properties and applications. Cambridge: Royal Society of Chemistry; 1999.

Russo RO, Berlyn GP. The use of organic biostimulants to help low input sustainable agriculture. J Sustain Agric. 1990 doi: 10.1300/J064v01n02_04. DOI

Tan KH. Humic matter in soil and the environment. 2. New York: Marcel Dekker; 2003.

Nardi S, Condheri G, Dell’Agnola G. Biological activity of humus. In: Piccolo A, editor. Humic substances in terrestrial ecosystems. Amsterdam: Elsevier; 1996. pp. 361–406.

Canellas LP, Olivares FL. Physiological responses to humic substances as plant growth promoter. Chem Biol Technol Agric. 2014 doi: 10.1186/2196-5641-1-3. DOI

Nardi S, Pizzeghello D, Schiavon M, Ertani A. Plant biostimulants: physiological responses induced by protein hydrolyzed-based products and humic substances in plant metabolism. Sci Agric. 2016 doi: 10.1590/0103-9016-2015-0006. DOI

Tejada M, Gonzalez JL. Influence of foliar fertilization with amino acids and humic acids on productivity and quality of asparagus. Biol Agric Hortic. 2003 doi: 10.1080/01448765.2003.9755270. DOI

Pizzeghello D, Nicolini G, Nardi S. Hormone-like activity of humic substances in Fagus sylvaticae forests. New Phytol. 2001 doi: 10.1046/j.0028-646x.2001.00223.x. PubMed DOI

Basiolio ZD, Pasqualoto CL, Facanha AR. Indolacetic and humic acids induce lateral root development through a concerted plasmalemma and tonoplast H+ pumps activation. Planta. 2007 doi: 10.1007/s00425-006-0454-2. PubMed DOI

Jackson TA, et al. Effects of clay minerals, oxyhydroxides, and humic matter on microbial communities of soil, sediment, and water. In: Huang PH, et al., editors. Environmental impact of soil component interactions: metals, inorganics and microbial activity. Berlin Heidelberg: CRC Press, Springer; 1995. pp. 165–200.

Parandian F, Samavat S. Effects of fulvic and humic acid on anthocyanin, soluble sugar, α-amylase enzyme and some micronutrient elements in Lilium. Int Res J Appl Basic Sci. 2012;3:924–929.

Dorer SP, Peacock CH. The effect of humate and organic fertilizer on establishment and nutrient of creeping bent putting greens. Int Turfgrass Soc. 1997;78:437–443.

Liu C, Cooper RJ, Bowman DC. Humic acid application affects photosynthesis, root development, and nutrient content of creeping bentgrass. HortScience. 1998;33:1023–1025. doi: 10.21273/HORTSCI.33.6.1023. DOI

Fernandez-Escobar R, Benlloch M. Response of olive trees to foliar application of humic substances extracted from leonardite. Sci Hortic. 1996 doi: 10.1016/S0304-4238(96)00914-4. DOI

Maibodi NDH, Kafi M, Nikbakht A, Rejali F. Effect of foliar applications of humic acid on growth, visual quality, nutrients content and root parameters of perennial ryegrass (Lolium perenne L.) J Plant Nutr. 2014 doi: 10.1080/01904167.2014.939759. DOI

Bettoni MM, Mogor AF, Pauletti V, Goicoechea N, Aranjuelo I, Germendia I. Nutritional quality and yield of onion as affected by different application methods and doses of humic substances. J Food Compos Anal. 2016 doi: 10.1016/j.jfca.2016.06.008. DOI

Olk DC, Dinnes DL, Rene Scoresby J, Callaway CR, Darlington JW. Humic products in agriculture: potential benefits and research challenges–a review. J Soils Sediments. 2018 doi: 10.1007/s11368-018-1916-4. DOI

Rose MT, Patti AF, Little KR, Brown AL, Roy Jackson W, Cavagnaro TR. A meta-analysis and review of plant-growth response to humic substances: practical implications for agriculture. Adv Agron. 2014 doi: 10.1016/B978-0-12-800138-7.00002-4. DOI

Lyons G, Genc Y. Commercial humates in agriculture: real substance or smoke and mirrors? Agronomy. 2016 doi: 10.3390/agronomy6040050. DOI

Kulikova NA, Abroskin DP, Badun G, Chernysheva MG, Korobkov VI, Beer AS, Tsvetkova EA, Senik SV, Klein OI, Perminova IV. Label distribution in tissues of wheat seedlings cultivated with tritium-labeled leonardite humic acid. Sci Rep. 2016 doi: 10.1038/srep28869. PubMed DOI PMC

Sedlacek P, Smilek J, Klucakova M. How the interactions with humic acids affect the mobility of ionic dyes in hydrogels—2. Non-stationary diffusion experiments. React Funct Polym. 2014 doi: 10.1016/j.reactfunctpolym.2013.12.002. DOI

Smilek J, Sedlacek P, Klucakova M. On the role of humic acids’ carboxyl groups in the binding of charged organic compounds. Chemosphere. 2015 doi: 10.1016/j.chemosphere.2015.06.093. PubMed DOI

Gladkov OA. U.S. Patent 7198805B2, Method for producing humic acid salts, Sankt-Peterburg (RU), chemical abstract (57). 2007.

Novak F, Sestauberova M, Hrabal R. Structural features of lignohumic acids. J Mol Struct. 2015 doi: 10.1016/j.molstruc.2015.03.054. DOI

Sedlacek P, Smilek J, Klucakova M. How the interactions with humic acids affect the mobility of ionic dyes in hydrogels—results from diffusion cells. React Funct Polym. 2013 doi: 10.1016/j.reactfunctpolym.2013.07.008. DOI

Sedlacek P, Smilek J, Kalina M, Lastuvkova M, Klucakova M. Hydrogels: invaluable experimental tool for demonstrating diffusion phenomena in physical chemistry laboratory courses. J Mater Educ. 2017;39:59–90.

Smilek J, Sedlacek P, Lastuvkova M, Kalina M, Klucakova M. Transport of organic compounds through porous systems containing humic acids. Bull Environ Contam Toxicol. 2017 doi: 10.1007/s00128-016-1926-0. PubMed DOI

Cussler EL. Diffusion: mass transfer in fluid systems. 2. London: Cambridge University Press; 1997.

Enev V, Pospisilova L, Klucakova M, Liptaj T, Doskocil L. Spectral characterization of selected humic substances. Soil Water Res. 2014 doi: 10.17221/39/2013-SWR. DOI

Prochazka P, Stranc P, Pzderu K, Strnc J, Jedlickova M. The possibilities of increasing the production abilities of soya vegetation by seed treatment with biologically active compounds. Plant Soil Environ. 2015 doi: 10.17221/225/2015-PSE. DOI

Prochazka P, Stranc P, Pazderu K, Stranc J. The influence of pre-sowing seed treatment by biologically active compounds on soybean seed quality and yield. Plant Soil Environ. 2016 doi: 10.17221/570/2016-PSE. DOI

Crank J. The mathematics of diffusion. 2. Oxford: Clarendon Press; 1956.

Guzman P, Fernandez V, Khayet M, Garcia ML, Fernandez A, Gil L. Ultrastructure of plant lead cuticles in relation to sample preparation as observed transmission electron microscopy. Sci World J. 2014 doi: 10.1155/2014/963921. PubMed DOI PMC

Tyree MT, Scherbatskoy D, Tabor CA. Leaf cuticles behave as asymmetric membranes: evidence from the measurement of diffusion potentials. Plant Physiol. 1990 doi: 10.1104/pp.92.1.103. PubMed DOI PMC

Guzman P, Fernandez V, Garcia ML, Khayet M, Fernandez A, Gil L. Localization of polysaccharides in isolated and intact cuticles of eucalypt, poplar and pear leaves by enzyme-gold labelling. Plant Physiol Biochem. 2014 doi: 10.1016/j.plaphy.2013.12.023. PubMed DOI

Guzman P, Fernandez V, Garcia J, Cabral V, Hayali N, Khayet M, Gil L. Chemical and structural analysis of Eucalyptus globulus and E. camaldulensis leaf cuticles: a lipidized cell wall region. Front Plant Sci. 2014 doi: 10.3389/fpls.2014.00481. PubMed DOI PMC

Fernandez V, Guzman-Delgado P, Graca J, Santos S, Gil L. Cuticles structure in relation to chemical composition: re-assessing the prevailing model. Front Plant Sci. 2016 doi: 10.3389/fpls.2016.00427. PubMed DOI PMC

Riederer M, Schönherr J. Accumulation and transport of (2,4-dichlorophenoxy)acetic acid in plant cuticles. 1. Sorption in the cuticular membrane and its components. Ecotoxicol Environ Saf. 1984 doi: 10.1016/0147-6513(85)90022-3. PubMed DOI

Eichert T, Burkhardt J. Quantification of stomatal uptake of ionic solutes using a new model system. J Exp Bot. 2001 doi: 10.1093/jexbot/52.357.771. PubMed DOI

Burkhardt J, Basi S, Pariyar S, Hunsche M. Stomatal penetration by aqueous solutions—an update involving leaf surface particles. New Phytol. 2012 doi: 10.1111/j.1469-8137.2012.04307.x. PubMed DOI

Schreiber L, Schonherr J. Water and solute permeability of plant cuticles. 1. Berlin: Springer; 2009.

Functional Hydrogels for Agricultural Application

How the Supramolecular Nature of Lignohumate Affects Its Diffusion in Agarose Hydrogel