CHEMICAL ENGINEERING TRANSACTIONS VOL. 79, 2020 A publication of The Italian Association of Chemical Engineering Online at www.cetjournal.it Guest Editors: Enrico Bardone, Antonio Marzocchella, Marco Bravi Copyright © 2020, AIDIC Servizi S.r.l. ISBN 978-88-95608-77-8; ISSN 2283-9216 Silver Nanoparticles Synthetized with Cinnamomum camphora to Reduces Total Coliforms in Soil Agricultural, Lima Perú Salumina Luz Beth Orizano Fabian, Elmer G. Benites Alfaro* César Vallejo University, Av. Alfredo Mendiola 6232 Lima 39, Perú. ebenitesa@ucv.edu.pe The objective of this work was to Determine the percentage of total coliform reduction by applying the silver nanoparticles synthesized with the Cinnamomum camphora substrate in the agricultural soil. First, the silver nanoparticles were synthesized being the reducing agent vegetal extract of Cinnamomum camphora, this also have antiseptic properties. The synthesis gave rise to silver nanoparticles with an average size of 37 nm. Second, it was the application of silver nanoparticles in four soil samples and in four different doses, which were 0.5 ml, 1.5 ml, 7 ml tond 10 ml and evaluated at different times. After the treatment and analysis of the total coliform concentration of the soil samples, to progressive decrease occurred ace time passed. After 72 hours, an average of 60% of the total coliform content in the agricultural soil was reduced, concluding that it is to viable way to improve the quality of the soils. However, other positive or negative impacts derived from the use of silver nanoparticles have not been yet evaluated; highlighting that the way in which nanoparticles have been synthesized is already an environmentally friendly method when not using chemical reagents. Keywords: Silver nanoparticles silver, synthesis whit green chemistry, reduction of coliforms in soils. 1. Introduction The problem of the pollution of the soil has done notorious in the last decade, since it sees affected by different types of contaminants, that are the product of the human activity, entering harmful substances like volatile organic compounds, toxic wastes of industry, domestic effluents, pesticides and fertilizers Like this, these substances affect gravely to the human health, vegetal and animal. A form to find a solution to the problems of pollution in the organisms receptors in the last times is the use of the nanotechnology. With the use of silver nanoparticles in an investigation obtained the reduction of 90% of bacteria in water, (Perez, 2011), in another case has allowed the control of the growth of bacteria lactic and acetic in the industry oenological, (García‐Ruiz To., et al., 2015). Also it has treated soil contaminated with TPHs (Fenantreno) using the nanoparticles metallic of elementary iron reducing between 22 to 50% (Valera, 2017). It has tested synthesis of suspensions of nanoparticles of copper and chitosan and tested his antimicrobic power in front of the Streptococcus mutans (Trepiana, 2015). On the other hand the obtaining of nanoparticles by means of vegetal extracts has come investigating by the environmental advantage that means, for example, for the application in paintings like additives in his function antifungal (Deyá and Bellotti, 2015). In other cases has tested the nanoparticles of silver to inhibit the training of biofilms of Pseudomona aeruginosa and Staphylococcus aureus, indicates that the bacterial capacity of the silver nanoparticles is high producing a mortality of until 99.9% (Flores C., 2014). The pollution of soil urge attention for being the natural reactor where develop multiple activities from processes of filtration, regulation of the cycle of water and biogeochemical, in addition to being universal habitat of living beings (Volke et al, 2004, pp.11-12). In a lot of countries does not exist rule related pathogens in soil, Peru is one of them, for this study took to theNo rma Mexican NOM-112-SSA1-1994, that establishes the permissible limits of microbiological characteristics in soil. The National Authority of the Water (ANA, 2016) has regulated the use of domestic residual waters treated in irrigation to avoid pollution of agricultural soil, since the pathogenic microorganisms are causers of damages to the health (Brennan J., DOI: 10.3303/CET2079055 Paper Received: 26 July 2019; Revised: 15 December 2019; Accepted: 23 February 2020 Please cite this article as: Orizano S., Benites E., 2020, Silver Nanoparticles Synthetized with Cinnamomum Camphora to Reduces Total Coliforms in Soil Agricultural, Lima Peru', Chemical Engineering Transactions, 79, 325-330 DOI:10.3303/CET2079055 325 2017). It has tested the use of copper, silver and zinc nanoparticles like nano fungicides in the treatment of pathogens (funguses) arrived from the soil to foliar plants obtaining that the copper nanoparticles were more efficient (Malandrakis A.A., 2019); however, it does not have still specific studies on the impacts of the nanoparticles in the soil for example in the biogeochemical cycle in the nitrogen cycle with the simultaneous application of fertilizers (Parada, J and et al, 2019). It has improved the quality of agricultural soil by means of the addition of seaweeds Litohothamnion calcareum nanoparticles for the improvement of production of fruit trees (Negreiros A.M.P., et al., 2019), in other cases has tested the improvement of mechanical stabilisation of soil with carbonate of calcium nanoparticles (Choobbasti J, et al, 2019). In this research, as in other previous works, Cinnamomum camphora extract polyphenols were used as reducing agents for synthetize silver nanoparticles. Then, these nanoparticles reduce the load of total coliforms in the soil. 2. Experimentation The following stages were followed: 2.1 Characterization of soil contaminated It determined the state of the physical chemistries and microbiological properties of the soil, specially the presence of total coliforms. The sample was collected following the Protocol of soils sampling (2014) of the Guide for Sampling of Soil, in concordance with the Supreme Decree N° 002-2017-MINAM (Standard of Environmental Quality - ECA for Soil) at Ministry of Environment (MINAM). 2.2 Syntheses of silver nanoparticles It used 20 g of leaves of Cinnamomum camphora previously wash, dried to acclimatise temperature. Next, it proceeded to make an extraction solid-liquid of the polyphenols of the Cinnamomum camphora using 3 g of this and crushing them, to a temperature of 60°C, in agitation to 900 rpm by a time of 20 minutes. Afterwards it filters, this extract was used for the synthesis later (Figure 1a). On the other hand, it prepared a solution of silver nitrate in deionized water taking 0.34 g of AgNO3 dissolving in 20 mL of water. For the synthesis stage, the AgNO3 solution is taken and with stirring of 900 rpm and at 60 ° C 4.4 mL of Cinnamomum camphora extract is added. When it presents a change of a brown colour it means will culminate the process of nanoparticles synthesis (Figure 1b), by means of the Tyndall effect (that it consists in observing the reflection of the light in the colloidal particles) verifies the nanoparticles presence since to simple sight are not visible. They were sent to a laboratory to verify their characterization. a b Figure 1a: Cinnamomum camphora Extract Figure 1b: Cinnamomum camphora Nanoparticles 2.3 Characterization of the nanoparticles The characterization of the nanoparticles did in the laboratory of sciences of the National University of Engineering, by means of dispersion of dynamic light (DLS), with the result that presents in the Graphic 1. It found nanoparticles of diameter of 37.2 nm. 326 Source: Laboratory of sciences-UNI Graphic 1: Size of nanoparticles of silver measured by DLS 2.4 Treatment of the soil with silver nanoparticles Silver nanoparticles synthesized in solution and stabilized at a pH around 7 using a buffer solution, were injected into the soil sample for a certain time. These were placed in four tanks where different amounts of silver nanoparticles were injected according to Figure 2. Sample of soil Dose of Ag nanoparticles pH 0.5 mL 6.8 1.5 mL 7.23 7 mL 7.45 10 mL 7.54 Source: Own Figure 2: Design of the samples of soil and dose of the nanoparticles used 2.5 Characterization of the soil afterwards of the treatment At this stage, the analysis was performed on soil samples treated with nanoparticles silver to determine the impact of the method related to the level total coliforms in the soil. These results are shown in the following section below. 3. Results and Discussion 3.1 State and characteristic initials of the soil The soil presented the physicochemical properties detailed in Table 1, highlighting that the soil was textural class, clay loam soil type and pH nearly to neutral. Analysis made in the Laboratory ANOBA LAB SAC. Initial Coliforms in the soil: The amount of total coliforms in the soil was 1,100 NMP / g on average, in the five soil monitoring points studied (Report of Laboratory ALAB Analytical E.I.R.L). M1 M2 M3 M4 327 Table 1: Physicochemical initial properties of the soil Physicochemical parameters Units Results Sand % 36 Silt % 30 Clay % 34 Class textural - clay loam PH (1/1) Und. pH 7.87 CE (1/1) dS/m 0.57 Carbonates %CaCO3 6.22 Organic matter oxidizable % 2.15 Interchangeable acidity meq/100g <0.2 Source: it Inform laboratory ANOBA LAB SAC 3.2 Total Coliforms in the soil after the treatment with silver nanoparticles (NPsAg) Considering the design shown in Figure 1, the soil treatment was made out with their respective repetitions to determine the level of total coliforms, in the times of 3, 48 and 72 hours after the start of the process. The analyses made in the Laboratory ALAB Analytical Laboratory E.I.R.L. using the norm of reference FDA/BAM, On-line 8th.ed.Rev.To, 3 1998. September 2002-Chapter 4, A, B, C, D, and F (Reviewed the 2013), con the Technician: Proofs of identification of coliforms organism: IMViC In three hours of treatment: In the M4 sample, the greatest level reduction of the total coliforms in the soil was obtained, in comparison to the other samples. It was reduced from 1,100 to 990 NMP/g equivalent to 10%, after treatment with NPsAg. See Graphic 2. Source: Own NMP (most likely number)٭ Graphic 2: Level of reduction of the total coliforms in the soil treated during 3 hours with solution containing silver nanoparticles. In 48 hours of treatment: Continuing the treatment with silver nanoparticles (NPsAg) in the same soil samples, at the end of 48 hours, it was found that in the M4 sample there was a greater reduction of total coliforms, coinciding that it is in this sample where also added greater amount of NPsAg, see Graphic 3a. In this sample it was reduced from 1,100 to 760 NMP/g, equivalent to 30% reduction of coliforms. In 72 hours of treatment: At 72 hours of treatment, in the M4 soil sample, was obtained the greatest reduction of total coliforms, 10 mL of NPsAg was used for the treatment and at pH of 7.54, (See Graphic 3b). The reduction was from 1,100 (initial) to 460 NMP/g, equivalent to 58.19%. 1,100 1,080 1,040 1,010 950 1000 1050 1100 1150 M0 M1 M2 M3 M4T ot al C ol ifo rm s (* N M P/ g) Sample number 3 hours 328 1,100 620 670 540 460 0 200 400 600 800 1000 1200 M0 M1 M2 M3 M4 To ta l C ol ifo rm s (N M P/ g) Sample number 72 hours a b Source: Own Source: Own Graphic 3a: Level of reduction of the total coliforms in the soil treated during 48 hours with solution containing silver nanoparticles. Graphic 3b: Level of reduction of the total coliforms in the soil treated during 72 hours with solution containing silver nanoparticles. This confirms that silver nanoparticles allow the reduction of total coliforms, as was investigated by Flores (2014), who reduced up to 99.9% of Pseudomona aeruginosa and Staphylococus aureus. Also for other pollutants, nanotechnology allows them to be reduced, such as Valera's research (2017) which, with elemental iron nanoparticles, achieved up to 50% reduction of phenanthrene. Therefore, the use of nanotechnology in the remediation of contaminated soil is an alternative, and there are already several projects carried out such as the European project NANOREM (2015), which after their tests confirm that it is possible to decontaminate soil and water effectively and safely. An important aspect of this topic is the synthesis of nanoparticles, the ecological form existing through the use of plant extracts as a reducing agent, for example, the research of Ledesma et al. (2014), among others. 4. Conclusion It was established that through the use of silver nanoparticles synthesized with Cinnamomum camphora extract of the size of 37 nm, it was able to reduce the total coliforms of a soil from the initial amount of 1,100 to 460 NMP/g. Therefore, it can be said that nanotechnology is the alternative way of using this practical and reliable method for the treatment of contaminated soils with a high coliform load. References Brennan J., 2017, The bacterium pathogenic in the soil. muyfitness.com/las-bacterias-patogenas-en-el- suelo_13071407/ Choobbasti A.J., Samakoosh M.A., Kutanaei S.S., 2019, Mechanical properties soil stabilized with nano calcium carbonate and reinforced with carpet waste fibers, Construction and Building Materials, 211, 1094–1104. doi.org/10.1016/j.conbuildmat.2019.03.306 Deyá C., Bellotti N., 2015, Vegetal extracts for the nanoparticles metallic synthesis and his application in paintings like antifungals additives, CIDEPINT (Investigation and Development Centre in Technology of Paintings)-CIC-CONICET, La Plata, Argentina. digital.cic.gba.gob.ar/bitstream/handle/11746/1356/t5-02.pdf-pdfa.pdf?sequence=1&isAllowed=and Flores C.Y., 2014, Silver Nanoparticles with potential applications on implantable materials: synthesis, characterization physical chemistry and bactericide activity, Thesis, La Plata University, Argentina. sedici.unlp.edu.ar/bitstream/handle/10915/34946/documento_completo.%20Flowers%20- %20Area%20Qu%C3%ADmica.Pdf?sequence=1 García‐Ruiz A., Crespo J, López‐of‐Luzuriaga J.M., Elms M., Monge M., Rodríguez‐Alfaro M.P., Martín‐ Álvarez P.J., Bartolome B. Moreno‐Arrive M.V., 2015, Synthesis and application of new silver nanoparticles biocompatibles for the control of the growth of lactics and acetics bacterium in wines. Rioja University. digital.csic.es/bitstream/10261/127113/1/nanopart%C3%ADculas%20of%20silver.Pdf 1,100 780 880 920 760 0 200 400 600 800 1000 1200 M0 M1 M2 M3 M4 To ta l C ol ifo rm s (N M P/ g) Sample number 48 hours 329 Ledezma A., Romero J., Hernández M., Moggio I., Arias E., Padrón G., Orozco V., Martínez A., Martínez C., Torres S., 2014, Synthesis biomimetic of silver nanoparticles using extract aqueous of nopal (Opuntia sp.) and his electro winning polymeric, Surfaces and empty, 27(4), pp. 133-140, Mexican Society of Science and Technology of Surfaces and Materials A.C. Federal district, Mexico. redalyc.org/pdf/942/94235742005.pdf Malandrakis A. A., Kavroulakis N., Chrysikopoulos C. V., 2019, Use of copper, silver and zinc nanoparticles against foliar and soil-borne plant pathogens, Science of the Total Environment, 670, 292–299. doi.org/10.1016/j.scitotenv.2019.03.210 Mexican Standard NOM-112-SSA1-1994, Determination of bacterium Coliform. Technique of the most probable number, Mexico, 1994. dof.gob.mx/nota_detalle.php?codigo=5398468&fecha=26/06/2015 MINAM, 2014, Guide of soil sampling. Environment Minister: Viceminister of Environmental quality, Lima Perú, p. 12. minam.gob.pe/wp-content/uploads/2014/04/guia-muestreo-suelo_minam1.pdf NANOREM, 2015, Nanotechnology for contaminated land remediation. nanorem.eu/projectdescription.aspx National authority of the Water–ANA, 2016, Manual of the best practices for the safe and productive use of the domestic residual water, Peru. ana.gob.pe/sites/default/files/publication/files/manual_de_buenas_practicas_para_el_uso_seguro_y_prod uctivo_de_las_aguas_residuales_domesticas.pdf Negreiros A.M.P., Sales R., Maia, F.F., Silva R.B., Costa J.A.P., Medeiros E.V. 2019, Lithothamnion calcareum Nanoparticles Increase Growth of Melon Plants, Notulae Botanicae Horti Agrobotanici Cluj- Napoca, 47(2), 426–431. doi.org/10.15835/nbha47111377 Parada J., Rubilar, O., Sousa D.Z., Martínez M., Fernández-Baldo M.A., Tortella, G.R., 2019, Short term changes in the abundance of nitrifying microorganisms in a soil-plant system simultaneously exposed to copper nanoparticles and atrazine, Science of the Total Environment, 670, p1068-1074. 7p. doi.org/10.1016/j.scitotenv.2019.03.221 Pérez S., 2011, Study of the microbial accumulation of metals and training of nanopartículas with potential application in the mining industry. Thesis, Institute Polytechnic National, México, p.5. epositoriodigital.ipn.mx/bitstream/123456789/15769/1/TesisSPB120112.pdf Trepiana D., 2015, Synthesis of suspensions of copper and quitosano nanoparticles and evaluation of his antimicrobic properties in front of Streptococcus mutans, Thesis, Chile University. repositorio.uchile.cl/bitstream/handle/2250/131959/s%C3%ADntesis-desuspensiones-of- nanopart%C3%ADculas-of-earn-and-quitosano-%20yevaluaci%C3%B3n-of-his-properties- antimicrobic.Pdf?sequence=1 Varela C., 2017, Evaluation of the effect of the nanoparticles metallic of elementary iron (NPHE) on the bacterium population of soil contaminated with TPHs (fenantreno) to laboratory level, Thesis, Ecuador Force Army University-ESPE. repositorio.espe.edu.ec/bitstream/21000/12877/1/t-espe-057176.pdf Volke T., Velasco J., De La Rosa D., s.f., Pollution by metals and metalloids: sampling and alternatives for his remediation, Mexico. books.google.com.pe/books?id=a50itx37scsc&printsec=frontcover&hl=is#v=on&epage q&f=true 330