Introduction
⌅Cocoa (Theobroma cacao L.) is a crop from South America and was brought to Central America in pre-Columbian times (Montemayor et al., 2002Montemayor JC, Risterucci AM, López PA, Ortiz CF, Moreno A, Lanaud C, 2002. Cacao domestication I: The origin of the cacao cultivated by the Mayas. Heredity 89(5): 380-386. https://doi.org/10.1038/sj.hdy.6800156). There are an estimated 6 to 7 million cocoa farmers worldwide, and 80-90% are smallholder farmers, with production areas between 2 and 4 ha, from tropical regions of Africa, Asia, Indonesia and Central and South America (Bermúdez et al., 2022Bermúdez S, Voora V, Larrea C, Luna E, 2022. Global Market Report. Cocoa prices and sustainability. https://www.iisd.org/publications?viewsreference[parent_entity_type]=node&viewsreference[parent_entity_id]=36&viewsreference[parent_field_name]=field_listing_type&page=10 [Jun 20, 2023].).
One of the aspects that affects cocoa cultivation commercialization is the presence of high concentrations of heavy metals (including Cd). This damages cocoa derivative quality, affecting 27% of exports (Huamaní-Yupanqui et al., 2012Huamaní-Yupanqui HA, Mansilla-Minaya LG, Florida-Rofner N, Neira-Trujillo GM, 2012. Presencia de metales pesados en cultivo de cacao (Theobroma cacao L.) orgánico. Acta Agronómica 61(4): 339-344.). Cd is generally adsorbed by roots in the soil and transported to the whole plant through the vascular system by essential cations, including Ca, Fe, and Zn (Huamaní-Yupanqui et al., 2012Huamaní-Yupanqui HA, Mansilla-Minaya LG, Florida-Rofner N, Neira-Trujillo GM, 2012. Presencia de metales pesados en cultivo de cacao (Theobroma cacao L.) orgánico. Acta Agronómica 61(4): 339-344.). High Cd concentrations in beans limit agricultural product marketing (Bravo Realpe et al., 2014Bravo Realpe IDS, Arboleda Pardo CA, Martín Peinado FJ, 2014. Efecto de la calidad de la materia orgánica asociada con el uso y manejo de suelos en retención de cadmio, en sistemas altoandinos de Colombia. Acta Agron 63(2): 164-174. https://doi.org/10.15446/acag.v63n2.39569).
Total Cd concentrations of geogenic origin in soil generally do not exceed 1 mg kg-1. However, in some cases levels of up to 16.3 mg kg-1 Cd have been found, which can be associated with weathering processes and the type of parental material, with a natural base level that would not exceed 0.5 mg kg-1 (Kabata-Pendias, 2010Kabata-Pendias A, 2010. Trace elements in soils and plants (4th ed). CRC PRESS, 548 pp. https://doi.org/10.1201/b10158).
There are many factors that influence Cd concentration in plants. Soil properties, such as soil starting material, soil organic matter (SOM), pH, carbonates, oxides and hydroxides of Fe and Mn, mineralogy of clays, salinity, anthropogenic contamination and plant genotype, affect the bioaccumulation and transfer of heavy metals to the cocoa bean (Alloway, 2013Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2). In turn, these factors can allow heavy metal dissolution, leaching and solubility (Alloway, 2013Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2).
The concentration of total Cd in the soil is not always a reliable indicator for predicting the concentration of Cd in the cocoa bean, as only one portion is bioavailable to the plant (Shahid et al., 2016Shahid M, Dumat C, Khalid S, Niazi NK, Antunes PMC, 2016. Cadmium bioavailability, uptake, toxicity and detoxification in soil plant system. Rev Environ Contam Toxicol 241: 73-137. https://doi.org/10.1007/398_2016_8). Alternatively, Cd and Zn in the soil present synergy when both metals have high concentrations; however, if one metal has higher concentration than the other, it is not absorbed by the plant (Huamaní-Yapanqui et al., 2012Huamaní-Yupanqui HA, Mansilla-Minaya LG, Florida-Rofner N, Neira-Trujillo GM, 2012. Presencia de metales pesados en cultivo de cacao (Theobroma cacao L.) orgánico. Acta Agronómica 61(4): 339-344.; Alloway, 2013Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2). The total Cd concentration in the soil can be transferred to the plant if the soil pH is very acidic or the SOM is low (Rábago & Aracil, 2011Rábago I, Aracil J, 2011. Buffer capacity for lead and cadmium contamination in soils in the Community of Madrid. PhD Thesis. Universidad Complutense, Madrid, Spain, 285 pp.).
The Cd in the plant is mobilized and transferred to the leaves and fruits, then concentrated in the grain, enabling us to verify the correlation between the bioavailable Cd in the soil and Cd in the grain (Argüello et al., 2019Argüello D, Chavez E, Lauryssen F, Vanderschueren R, Smolders E, Montalvo D, 2019. Soil properties and agronomic factors affecting cadmium concentrations in cacao beans: A nationwide survey in Ecuador. Sci Total Environ 649: 120-127. https://doi.org/10.1016/j.scitotenv.2018.08.292; Engbersen et al., 2019Engbersen N, Gramlich A, López M, Schwarz G, Hattendorf B, Gutiérrez O, et al., 2019. Cadmium accumulation and allocation in different cacao cultivars. Sci Total Environ 678: 660-670. https://doi.org/10.1016/j.scitotenv.2019.05.001).
A high intake of cocoa beans at the maximum concentrations permitted by the Codex Alimentarius (FAO/WHO, 2017FAO/WHO, 2017. Codex Alimentarius (Codex). Report of the 11th meeting of the Codex Committee on Food Contaminants. http://www.fao.org/fao-who-codexalimentarius/codex-texts/guidelines/en/. [May 29, 2021].) in cocoa bean derivatives may promote the accumulation of Cd in the human body. This may cause serious problems in tissues and organs that are susceptible to it, such as the kidneys and liver (Casteblanco, 2018Casteblanco JA, 2018. Heavy metals remediation techniques with potential application in cocoa cultivation. La Granja 27(1): 21-35. https://doi.org/10.17163/lgr.n27.2018.02).
Furcal-Beriguete & Torres-Morales (2020)Furcal-Beriguete P, Torres-Morales JL, 2020. Determinación de concentraciones de cadmio en plantaciones de Theobroma cacao L. en Costa Rica. Revista Tecnología En Marcha 33(1): 122-137. https://doi.org/10.18845/tm.v33i1.5027 report that the European Food Safety Authority has not established a limit on Cd concentration in the unfermented or raw bean, but there are studies that show that Cd concentrations in the fermented and dried cocoa bean reach levels higher than established. The maximum limit for chocolate and other products with cocoa content between 50 and 70% is 0.6 mg kg-1, while chocolates and products with declared cocoa content above 70% the maximum level is 0.8 mg kg-1.
Cocoa cultivation in Panama presents high Cd concentrations in the bean, reducing its quality, price and marketing possibilities. Unlike other countries such as Peru (Huamaní-Yupanquí et al., 2012Huamaní-Yupanqui HA, Mansilla-Minaya LG, Florida-Rofner N, Neira-Trujillo GM, 2012. Presencia de metales pesados en cultivo de cacao (Theobroma cacao L.) orgánico. Acta Agronómica 61(4): 339-344.), Costa Rica (Florida, 2021Florida Rofner N, 2021. Review of maximum limits for cadmium in cocoa (Theobroma cacao L.). La Granja 34(2): 117-130. https://doi.org/10.17163/lgr.n34.2021.08) and Ecuador (Barraza et al., 2017Barraza F, Schreck E, Lévêque T, Uzu G, López F, Ruales J, et al., 2017. Cadmium bioaccumulation and gastric bioaccessibility in cacao: A field study in areas impacted by oil activities in Ecuador. Environ Pollut 229: 950-963. https://doi.org/10.1016/j.envpol.2017.07.080), in Panama there are no studies on the relationships between soil Cd and its presence in cocoa beans, despite the fact that moderate concentrations of the metal are usually detected (personal communication with members of the Cooperative of multiple services of cacao Bocatoreño R.L).
It is pertinent to note that certain agronomic management such as the application of organic fertilizers, biochar, beneficial microorganisms and liming can also reduce the absorption of Cd and other heavy metals in cocoa (Dávila-Zamora et al., 2020Dávila-Zamora EC, Reyes-Evangelista LA, Aldoradin-Puza E, Londoñe-Bailon P, Aleman-Polo M, 2020. Cd and Pb reduction in cocoa (Theobroma cacao L.) nib using two organic amendments. Revista Colombiana de Investigaciones Agroindustriales 7(1): 20-29. https://doi.org/10.23850/24220582.2776). But cocoa cultivation areas in Panama do not maintain an adequate fertilization program.
This paper evaluates the concentration of Cd in cocoa plants, especially in the bean, and its relationship with the available Cd and other soil properties of small producer’s fields in the province of Bocas del Toro, Panama. Our objective was to calculate Cd concentration in cocoa plants and evaluate its relationship with available Cd and other soil properties.
Material and methods
⌅Study site location
⌅The research was conducted during 2020-2021 in the district of Almirante, province of Bocas del Toro, Republic of Panama, on 21 farms cultivated with cocoa of various genotypes within agroforestry systems (Fig.1).
The climate of the province of Bocas del Toro is classified as very humid tropical. Annual precipitation is 2735 mm with an average annual temperature of 24.3 °C (Peel et al., 2007Peel MC, Finlayson BL, McMahon TA, 2007. Updated world map of the Köppen-Geiger climate classification. Hydrol Earth Syst Sci 11(5): 1633-1644. https://doi.org/10.5194/hess-11-1633-2007). The soils of the district of Almirante are classified taxonomically within the order of Inceptisol (Soil Survey Staff, 2014Soil Survey Staff, 2014. Keys to Soil Taxonomy, 12th Edition, USDA-Natural Resources Conservation Service, Washington DC.).
Field work and sample preparation
⌅The study was carried out in eight producing farms of the district of Almirante, and within each farm two or three sampling plots were selected depending on the area of the plantation (SEMARNAT, 2002SEMARNAT, 2002. Official Mexican Standard NOM-021-RECNAT-2000 that establishes the specifications of fertility, salinity and classification of soils, study, sampling and analysis. Secretary of Environment and Natural Resources, Mexico, 85 pp.) to complete a total of 21 plots. Each plot of 300 m2 (10 × 30 m) was located at representative sites of the plantation.
In each of these plots, 15 soil subsamples were systematically taken in the form of zig-zags equidistant from each other at a distance of approximately 1.5 m from the trunk of each cocoa tree. The soil samples were taken up to 30 cm deep, and with the 15 subsamples a homogenized composite sample of approximately 2 kg was made.
Leaf and fruit sampling was carried out within 300 m2 containing 17 cocoa trees established in a planting frame of 3 × 3 m. Fifty-one healthy leaves were collected, three for each tree, located in the middle part of the cocoa tree, taking the fourth leaf from the apex of the selected branch following the methodology of Puentes et al. (2016)Puentes PYJ, Menjivar JC, Aranzazu F, 2016. Concentration of nutrients in leaves, a tool for nutritional diagnosis in cocoa. Agron Mesoam 27(2): 329-336. https://doi.org/10.15517/am.v27i2.19728. Similarly, six fruits of uniform size were collected within the selected area (300 m2) systematically (1 fruit for every 3 plants).
In the laboratory, rootlets, stones or any material that could alter soil analysis were removed. Then the soil samples were air-dried and crushed in an Agatha mortar and passed through a 2-mm nylon mesh sieve for all analyses except Cd, where another 0.063 mm sieve was also used according to the Soil Sampling Guide of the Ministry of the Environment, Peru (MINAM, 2014MINAM, 2014. Guía para el muestreo de suelo. En el marco del Decreto Supremo Nº.002-2013-MINAM, Estándares de Calidad Ambiental (ECA) para suelo. Ministerio del Ambiente, Perú.).
On the other hand, the leaves and fruits were washed with doubly distilled water (type 2) eliminating the soil particles adhered to them. The leaves were taken to the stove for drying at a temperature of 60 °C until a constant weight was maintained. For the fruit, the cocoa beans plus mucilage were separated from the rest of the fruit. Both parts were dried, following the same procedure as leaves. Finally, the leaves, fruits and grains were crushed.
Soil analysis
⌅The pH was determined by potentiometry with a ratio of 1:2.5 (soil-water) using techniques described by Teixeira et al. (2017)Teixeira PC, Donagemma GK, Fontana To, Teixeira WG, 2017. Manual de métodos de análise do solo, 3rd ed. rev. Brasília, DF: Embrapa, 573 pp.. Soil organic carbon (SOC) was analyzed using the method of Walkley & Black (1934)Walkley A, Black AI, 1934. An examination of the method for determination soil organic matter, and a proposed codification of the chromic acid titration method. Soil Sci 37: 29-38. https://doi.org/10.1097/00010694-193401000-00003. Total N was determined by Kjeldahl digestion following the methodologies of Teixeira et al. (2017)Teixeira PC, Donagemma GK, Fontana To, Teixeira WG, 2017. Manual de métodos de análise do solo, 3rd ed. rev. Brasília, DF: Embrapa, 573 pp..
For the extraction of bioavailable elements (Cu, Fe, Zn, Mn, Cd) and cations such as Ca, Mg and K, the Mehlich 3 extractor solution (Mehlich, 1984Mehlich A, 1984. Mehlich 3 soil test extractant: A modification of Mehlich 2 extractant. Commun Soil Sci Plant Anal 15(12): 1409-1416. https://doi.org/10.1080/00103628409367568) was used, taking a soil: water ratio 1:10 (w/v), and the analysis by atomic absorption spectrophotometer of continuous source of high-resolution flame techniques (FAAS) was subsequently performed.
The exchangeable Al was extracted with 1 M KCl extractor solution, using techniques described by Teixeira et al. (2017)Teixeira PC, Donagemma GK, Fontana To, Teixeira WG, 2017. Manual de métodos de análise do solo, 3rd ed. rev. Brasília, DF: Embrapa, 573 pp.. The effective cation exchange capacity (ECEC) was determined by the sum of the bases (Ca, Mg, K) + Al. Aluminium’s saturation percentage was determined as follows: Al. Sat. (%) = Al / ECEC * 100. The granulometric physical analysis of the soil texture was carried out using the methodology manual (Teixeira et al., 2017Teixeira PC, Donagemma GK, Fontana To, Teixeira WG, 2017. Manual de métodos de análise do solo, 3rd ed. rev. Brasília, DF: Embrapa, 573 pp.).
The total concentrations of Cu, Zn, Fe, Mn, Cd, K, Ca and Mg in soil, leaves, fruits and cocoa beans were evaluated by microwave-assisted extraction using reagents with pure concentrations of HNO3 and HCl in a 3:1 ratio and subsequently analyzed by high-resolution continuous source atomic absorption spectrophotometry of flame techniques, according to USEPA 3051A methodologies (USEPA, 2007USEPA, 2007. Method 3051A (SW-846): Microwave assisted acid digestion of sediments, sludges, and oils. Revision 1. Washington, DC.). With this methodology we calculated the limits of detection (LOD), the limits of quantification (LOQ) and the adjusted R2 for the elements evaluated.
Statistical analysis
⌅Summary statistics and confidence intervals for estimators were used to describe the data. Pearson linear correlation analyses were performed to determine the association between soil variables and Cd in the plant.
To take into account the joint effect of the physicochemical variables of the soil on the concentrations of available Cd, a multiple-linear regression analysis was conducted. This same analysis was performed to evaluate how bioavailable Cd and other physicochemical soil variables relate to Cd in plants (leaves, fruits and grains). In multiple regressions, the backward variable selection method was used, retaining significant regressors (p<0.05) and to avoid multicollinearity, values of inflation of variance (VIF) less than five were considered (Balzarini et al., 2008Balzarini MG, González L, Tablada M, Casanoves F, Di Rienzo JA, Robledo CW, 2008. Infostat. User Manual, Editorial Brujas, Córdoba, Argentina. 331 pp.). The model assumptions were evaluated by means of a graphic inspection of the residues. The analyses were performed using the statistical software InfoStat version 2020 (Di Rienzo et al., 2020Di Rienzo JA, Casanoves F, Balzarini MG, González L, Tablada M, Robledo CW, 2020. InfoStat vers 2020. Centro de Transferencia InfoStad, FCA, Universidad Nacional de Córdoba, Argentina. URL http://www.infostat.com.ar.).
Results
⌅In Table 1, we present the limits of detection and instrumental quantification for the eight elements quantified on the atomic absorption spectrophotometer equipment, which met the requirements for precision and accuracy. By using it, the results were obtained with the necessary level of reliability. Additionally, summary measurements and 95% confidence intervals were used for soil mineral content and physical properties extracted at a depth of 30 cm from 21 evaluated plots (Table 2).
| Element | Unit | LOD | LOQ | R2 (adj) |
|---|---|---|---|---|
| K | cmol(+) kg-1 | 0.037 | 0.112 | 0.996 |
| Ca | cmol(+) kg-1 | 0.004 | 0.012 | 0.969 |
| Mg | cmol(+) kg-1 | 0.128 | 0.383 | 0.992 |
| Cu | mg kg-1 | 0.0031 | 0.093 | 0.999 |
| Zn | mg kg-1 | 0.001 | 0.002 | 0.996 |
| Fe | mg kg-1 | 0.007 | 0.022 | 0.994 |
| Mn | mg kg-1 | 0.003 | 0.008 | 0.984 |
| Cd | mg kg-1 | 0.001 | 0.002 | 0.999 |
The soils showed high acidity, with a mean pH of 5.1 and a 95% confidence interval of 4.9-5.3. SOM levels were moderate, at 5.1%. There was an average concentration of 0.34 % of total nitrogen, with minimum and maximum values of 0.1 and 0.6 %, respectively. Among the sites sampled for the C: N ratio, the mean value was 9.9, with minimum levels of 4.5 and maximum levels of 26.
A value in the mean of the ECEC of 19.9 cmol(+) kg-1 (Table 2) was observed, this level being high in Inceptisol soils. In the same way, the elements that are part of the ECEC, K, Ca and Mg, obtained high concentrations with means of 0.20, 14.1 and 5.0 cmol(+) kg-1, respectively. The average concentrations of exchangeable Al had a moderate value of 0.7 cmol(+) kg-1, with a low level of Al. Sat. 3.9%, being able to find in the plots, minimum and maximum concentrations of 0.6 and 13.9%, respectively.
| Variable | Unit | Mean | SE | CV | Min | Max | Median | LI | UL |
|---|---|---|---|---|---|---|---|---|---|
| 95% | |||||||||
| pH (H2O) | 5.10 | 0.06 | 7.60 | 4.40 | 6.00 | 5.10 | 4.90 | 5.30 | |
| SOM | % | 5.10 | 0.25 | 31.70 | 1.80 | 10.10 | 4.90 | 4.50 | 5.80 |
| SOC | % | 2.90 | 0.15 | 31.70 | 1.10 | 5.90 | 2.90 | 2.60 | 3.40 |
| Total N | % | 0.34 | 0.02 | 37.10 | 0.10 | 0.60 | 0.40 | 0.30 | 0.40 |
| Ratio (C:N) | 9.90 | 0.64 | 41.80 | 4.50 | 26.10 | 9.10 | 8.20 | 11.60 | |
| ECEC | cmol(+) kg-1 | 19.90 | 1.33 | 43.30 | 6.40 | 38.70 | 17.60 | 15.90 | 23.90 |
| K | cmol(+) kg-1 | 0.20 | 0.02 | 69.90 | 0.04 | 0.60 | 0.14 | 0.14 | 0.30 |
| Ca | cmol(+) kg-1 | 14.10 | 0.97 | 44.70 | 3.20 | 28.30 | 11.40 | 11.20 | 16.90 |
| Mg | cmol(+) kg-1 | 5.00 | 0.41 | 52.60 | 1.50 | 11.10 | 4.60 | 3.80 | 6.20 |
| Al | cmol(+) kg-1 | 0.70 | 0.10 | 97.20 | 0.10 | 2.50 | 0.40 | 0.40 | 0.90 |
| Al. Sat. | % | 3.90 | 0.56 | 96.00 | 0.60 | 13.90 | 2.90 | 2.10 | 5.50 |
| Cu | mg kg-1 | 2.50 | 0.26 | 68.60 | 0.00 | 5.90 | 2.50 | 1.70 | 3.30 |
| Zn | mg kg-1 | 3.60 | 0.40 | 70.70 | 0.60 | 10.90 | 2.40 | 2.50 | 4.80 |
| Fe | mg kg-1 | 101.90 | 8.20 | 52.30 | 49.50 | 250.20 | 86.40 | 77.80 | 12.10 |
| Mn | mg kg-1 | 177.10 | 16.40 | 59.90 | 50.00 | 491.30 | 147.90 | 128.40 | 225.80 |
| Total Cd | mg kg-1 | 0.10 | 0.02 | 36.20 | 0.10 | 0.12 | 0.10 | 0.10 | 0.14 |
| Cdbd | mg kg-1 | 0.02 | 0.01 | 32.20 | 0.02 | 0.03 | 0.02 | 0.02 | 0.03 |
| Sand | % | 57.80 | 2.04 | 22.80 | 36.00 | 76.00 | 60.00 | 52.00 | 63.60 |
| Silt | % | 28.90 | 1.72 | 38.40 | 12.00 | 48.00 | 27.50 | 24.10 | 33.90 |
| Clay | % | 13.30 | 1.01 | 49.00 | 1.00 | 24.00 | 13.50 | 10.60 | 16.30 |
SE: standard error. CV: coefficient of variation. LI: lower limit. UL: upper limit. SOM: soil organic matter. SOC: soil organic carbon. ECEC: effective cation exchange capacity. Al. Sat.: aluminum saturation. Cdbd: bioavailable cadmium.
The micronutrients in the soil presented, for the concentration of Cu, Zn, Fe and Mn extracted by Mehlich 3, values of 2.5, 3.6, 101.9 and 177.1 mg kg-1, respectively. Soil concentrations of total Cd and bioavailable Cd (Cdbd) yielded mean values of 0.10 and 0.02 mg kg-1, respectively, whose 95% confidence intervals were 0.07-0.14 and 0.02-0.03 mg kg-1, respectively.
With respect to soil texture, the largest fraction corresponded to sand particles (57.8%), followed by silt (28.9%) and clay (13.3%).
A description of the Cd concentration in cocoa leaves, fruits, and beans is presented in Table 3 as well as 95% confidence intervals. A minimum and maximum limit is also provided in Fig. 2. The average concentration of Cd in leaves (Cdleaves) was found to be 1.04 mg kg-1 with a minimum and maximum concentration of 0.13-4.40 mg kg-1. The concentration of Cd in fruits (Cdfruits) was 0.34 mg kg-1 with a minimum concentration of 0.09 mg kg-1 and maximum of 0.76 mg kg-1. The concentration of Cd in cocoa beans (Cdbeans) was 0.25 mg kg-1, with minimum and maximum levels of 0.01-0.95 mg kg-1 respectively with lower limits of 0.15 mg kg-1 and upper limits of 0.34 mg kg-1.
| Variable1 | Mean | SE | CV | Min | Max | Median | LI | UL |
|---|---|---|---|---|---|---|---|---|
| 95% | ||||||||
| Cdleaves | 1.04 | 0.15 | 90.9 | 0.13 | 4.40 | 0.74 | 0.60 | 1.40 |
| Cdfruits | 0.34 | 0.03 | 52.3 | 0.09 | 0.76 | 0.30 | 0.30 | 0.40 |
| Cdbeans | 0.25 | 0.03 | 90.7 | 0.01 | 0.95 | 0.14 | 0.20 | 0.30 |
SE: standard error. CV: coefficient of variation. LI: lower limit. UL: upper limit. Cdleaves: cadmium in leaves. Cdfruits: cadmium in fruits. Cdbeans: cadmium in beans.
Pearson’s correlations were calculated to evaluate the relationship between Cdbeans concentration and soil variables (Table 4). Pearson’s correlations between the concentration variables of Cdbd, Cdleaves and Cdfruits were also calculated (Table 5).
| Variables | R | p-value | |
|---|---|---|---|
| Cdbeans | Cdbd | 0.50 | 0.0243 |
| Cdbeans | Total Cd | 0.66 | 0.0010 |
| Cdbeans | SOM (%) | -0.50 | 0.0207 |
| Cdbeans | Ratio (C:N) | 0.05 | 0.8432 |
| Cdbeans | Silt (%) | 0.47 | 0.0316 |
| Cdbeans | Clay (%) | 0.37 | 0.1005 |
| Cdbeans | pH | -0.04 | 0.8773 |
| Cdbeans | Al | 0.36 | 0.1138 |
| Cdbeans | Casoil | -0.06 | 0.8114 |
| Cdbeans | Mgsoil | -0.12 | 0.5947 |
| Cdbeans | Ksoil | -0.23 | 0.3052 |
| Cdbeans | ECEC | -0.05 | 0.8138 |
| Cdbeans | Cusoil | -0.17 | 0.4634 |
| Cdbeans | Znsoil | -0.38 | 0.0929 |
| Cdbeans | Fesoil | 0.47 | 0.0317 |
| Cdbeans | Mnsoil | -0.14 | 0.5391 |
Cddb: bioavailable cadmium. SOM: soil organic matter. ECEC: effective cation exchange capacity. Bold probabilities indicate significant correlation (p<0.05).
| Variables | R | p-value | |
|---|---|---|---|
| Cdbd | Cdleaves | 0.35 | 0.1169 |
| Cdbd | Cdfruits | 0.49 | 0.0234 |
| Cdleaves | Cdfruits | 0.57 | 0.0075 |
| Cdleaves | Cdbeans | 0.84 | <0.0001 |
| Cdfruits | Cdbeans | 0.64 | 0.0016 |
Cdbd: bioavailable cadmium. Bold probabilities indicate significant correlation (p<0.05).
Cdbeans (Table 4) was positively correlated with total Cd and Cdbd (r = 0.66 and r = 0.50 respectively; p<0.05). A positive correlation was also found with the percentage of slime and the content of Fe (r = 0.47 in both; p<0.05). The content of organic matter and Zn maintained a negative correlation with Cd in the cocoa bean with values of (r = -0.50; p<0.05) and (r = -0.38; p<0.10) respectively.
There was no significant correlation between Cdleaves and Cbd (p=0.1169, Table 5). The concentration of Cdleaves presented a positive correlation with Cdbeans (r = 0.84, p<0.0001) and with Cdfruits (r = 0.57, p = 0.0075). In turn, the Cdfruits correlate positively with the Cdbeans (r = 0.64, p=0.0016).
The relationship between the concentration of bioavailable Cd (Cdbd) and the physicochemical characteristics of the soil could be modeled with five variables according to the backward selection method (Table 6a ), where the adjusted R2 of the multiple regression model was 0.54. The resulting model was: Cdbd = 0.16 + 5.4 × 10-4 Silt (%) - 8.9 × 10-4 Clay (%) - 0.03 pH + 1.9 × 10-3 Zn + 1.2 × 10-4 Fe.
| Variable | Unit | Estimate | SE | LI | UL | T | p-value | CP Mallows | FIV |
|---|---|---|---|---|---|---|---|---|---|
| 95% | |||||||||
| a) Bioavailable Cd | |||||||||
| Constant | -- | 0.16 | 0.03 | 0.09 | 0.22 | 4.97 | 0.0002 | ||
| Silt | % | 5.4·10-4 | 1.60·10-4 | 2·10-4 | 8.8·10-4 | 3.35 | 0.0044 | 15.24 | 1.64 |
| Clay | % | -8.9·10-4 | 2.60·10-4 | -1.4·10-3 | -3.4·10-4 | -3.44 | 0.0036 | 15.84 | 1.44 |
| pH | -- | -0.03 | 0.01 | -0.05 | -0.02 | -4.53 | 0.0004 | 24.53 | 3.56 |
| Zn | mg kg-1 | 1.9·10-3 | 6.7·10-4 | 4.7·10-4 | 3.3·10-4 | 2.84 | 0.0124 | 12.06 | 1.64 |
| Fe | mg kg-1 | 1.2·10-4 | 4.3·10-5 | 3·10-4 | 2.1·10-4 | 2.83 | 0.0126 | 12.02 | 2.81 |
| b) Cd in cocoa leaves | |||||||||
| Constant | -- | -0.26 | 0.62 | -1.51 | 1.00 | -0.41 | 0.6810 | ||
| Al | cmol (+) kg-1 | 0.50 | 0.18 | 0.13 | 0.86 | 2.78 | 0.0085 | 11.74 | 1.15 |
| Silt | % | 0.02 | 0.01 | 3.2·10-3 | 0.05 | 2.32 | 0.0259 | 9.40 | 1.25 |
| Clay | % | 0.04 | 0.02 | 3.2·10-3 | 0.07 | 2.21 | 0.0337 | 8.87 | 1.17 |
| SOM | % | -0.15 | 0.07 | -0.29 | -2.7·10-3 | -2.07 | 0.0461 | 8.27 | 1.15 |
| Cdbd | mg kg-1 | 22.37 | 10.62 | 0.82 | 43.91 | 2.11 | 0.0423 | 8.43 | 1.11 |
| c) Cd in cocoa fruits | |||||||||
| Constant | -- | 0.36 | 0.10 | 0.15 | 0.57 | 3.67 | 0.0021 | ||
| Cdbd | mg kg-1 | 11.4 | 3.19 | 4.63 | 18.2 | 3.57 | 0.0026 | 15.75 | 1.06 |
| Al | cmol (+) kg-1 | -0.09 | 0.05 | -0.20 | 0.01 | -1.97 | 0.0669 | 6.86 | 1.27 |
| K | cmol (+) kg-1 | -0.47 | 0.21 | 0.92 | -0.02 | -2.21 | 0.0423 | 7.87 | 1.17 |
| Zn | mg kg-1 | -0.03 | 0.01 | -0.05 | -3.8·10-3 | -2.44 | 0.0265 | 8.97 | 1.17 |
| d) Cd in cocoa beans | |||||||||
| Constant | -- | -0.64 | 0.2 | -1.06 | -0.21 | -3.19 | 0.0057 | ||
| Cdbd | mg kg-1 | 20.44 | 3.6 | 12.81 | 28.07 | 5.68 | >0.0001 | 35.23 | 1.39 |
| Zn | mg kg-1 | -0.03 | 0.01 | -0.05 | -2.7·10-3 | -2.36 | 0.0313 | 8.57 | 1.11 |
| Clay | % | 0.03 | 0.01 | 0.01 | 0.04 | 4.45 | 0.0006 | 21.01 | 1.87 |
| Mg | cmol (+) kg-1 | 0.04 | 0.01 | 0.01 | 0.07 | 2.91 | 0.0103 | 11.46 | 1.89 |
SE: standard error. LI: lower limit. UL: upper limit. T: T-statistic. CP: predictive coefficient. FIV: values of inflation of variance. SOM: soil organic matter. Cdbd: bioavailable cadmium.
The concentration of Cdbd is mostly influenced by soil pH, due to the high predictive coefficient (Cp) of Mallows (24.53); the lower the soil pH there is more Cdbd. The percentage content of clay follows in predictive value to the pH; the lower the percentage of clay, the higher the concentration of Cdbd. The opposite occurs with the percentage of silt, the higher the percentage more Cdbd. Zn and Fe complete the set of significant variables to explain Cdbd; in this case the higher the Fe and Zn, the higher the content of Cdbd.
The relationship between cadmium in leaves (Cdleaves) and the physicochemical characteristics of the soil could be modeled with five variables according to the backward method (Table 6b ). The adjusted R2 of the model was 0.47.
The estimated multiple regression model was: Cdleaves = - 0.26 + 0.50 Al + 0.02 Silt (%) + 0.04 Clay (%) - 0.15 SOM (%) + 22.37 Cdbd, interpreting that the variables that better predict according to Mallows’ Cp were exchangeable Al (11.74) and silt content (9.40), while Cdbd had a positive relationship with Cdleaves.
The relationship between Cdfruits and the physicochemical characteristics of the soil could be modeled with fiur variables according to the backward method (Table 6c). The adjusted R2 of the model was 0.44. The resulting model for Cd in fruits was: Cdfruits = 0.36 + 11.40 Cdbd - 0.09 Al -0.47 K - 0.03 Zn.
According to Mallows’ Cp, the variables that better predict Cd concentration in fruits were Cdbd (15.75) and Zn (8.97). Except for Cdbd, which had a positive relationship with Cdfruits, the other variables affected it negatively.
The relationship between the concentration of Cdbeans and the physicochemical characteristics of the soil was modeled with four variables according to the backward method (Table 6d). The adjusted R2 of the model was 0.68. The resulting model was: Cdbeans= - 0.64 + 20.44 Cdbd - 0.03 Zn + 0.03 Clay (%) + 0.04 Mg.
As in the case of Cdfruits, the variable of greater intervention for the concentration of Cdbeans was the Cdbd with a Cp of Mallows of 35.23 and with a positive association, as well as Mg and Clay. The only variable negatively affecting the amount of Cdbeans was the Zn content.
Discussion
⌅In soil, the concentration of heavy metals (e.g., Cd) is closely linked to the content of SOM, which forms organometallic complexes, immobilizing their absorption by plants (Arévalo-Gardini et al., 2016Arévalo-Gardini E, Obando-Cerpa ME, Zúñiga-Cernades LB, Arévalo-Hernández CO, Baligar VC, He ZL, 2016. Heavy metals in soils of cocoa plantations (Theobroma cacao L.) in three regions of Peru. Appl Ecol 15(2): 81-89. https://doi.org/10.21704/rea.v15i2.747). Researchers such as Cameselle & Gouveia (2019)Cameselle C, Gouveia S, 2019. Phytoremediation of mixed contaminated soil enhanced with electric current. J Hazard Mater 361: 95-102. https://doi.org/10.1016/j.jhazmat.2018.08.062 mention that bioavailable Cd in soil can vary due to interactions with other metals. In turn, the concentration of Cd in the soil depends on the depth at which the sample is taken (Goebes et al., 2019Goebes P, Schmidt K, Seitz S, Both S, Bruelheide H, Erfmeier A, et al., 2019. The strength of soil-plant interactions under forest is related to a critical soil depth. Sci Rep 9: 8635. https://doi.org/10.1038/s41598-019-45156-5).
The average pH value found in this study is considered very acidic (Snoeck et al., 2016Snoeck D, Koko L, Joffre J, Bastide P, Jagoret P, 2016. Cocoa nutrition and fertilization. In: Sustainable agriculture reviews, vol 19; Lichtfouse E (ed). Springer International Publishing, Cham. pp: 155-202. https://doi.org/10.1007/978-3-319-26777-7_4). Low pH values allow for the highest solubility of toxic elements such as Al (0.7 cmol(+) kg-1 on average in this study) and Cd. This is significant in cocoa cultivation since the absorption processes of the Cd concentration are favored when the pH is very acid. In the same way, when the pH in the soil increases, Cd is adsorbed by colloids, blocking its bioavailability (Liu et al., 2020Liu N, Jiang Z, Li X, Liu H, Li N, Wei S, 2020. Mitigation of cadmium (Cd) accumulation in rice through the joint application of organic amendments and selenium (Se) in contaminated soils with high Cd content. Chemosphere 241: 125106. https://doi.org/10.1016/j.chemosphere.2019.125106).
This could be verified in the regression model (Table 6a) where the pH of the soil was presented with a p-value < 0.05 and a high prediction value, indicating that is the variable that most interferes in the concentration of Cdbd; the lower the pH, the more Cdbd. Several studies affirm that the bioavailability of Cd is closely related to pH values (Arévalo-Gardini et al., 2016Arévalo-Gardini E, Obando-Cerpa ME, Zúñiga-Cernades LB, Arévalo-Hernández CO, Baligar VC, He ZL, 2016. Heavy metals in soils of cocoa plantations (Theobroma cacao L.) in three regions of Peru. Appl Ecol 15(2): 81-89. https://doi.org/10.21704/rea.v15i2.747; Florida, 2021Florida Rofner N, 2021. Review of maximum limits for cadmium in cocoa (Theobroma cacao L.). La Granja 34(2): 117-130. https://doi.org/10.17163/lgr.n34.2021.08).
Generally, the average SOM content is greater than 3% (Table 2) which is adequate for cocoa cultivation (Snoeck et al., 2016Snoeck D, Koko L, Joffre J, Bastide P, Jagoret P, 2016. Cocoa nutrition and fertilization. In: Sustainable agriculture reviews, vol 19; Lichtfouse E (ed). Springer International Publishing, Cham. pp: 155-202. https://doi.org/10.1007/978-3-319-26777-7_4) while helping the nutrition of the crop and immobilization of bioavailable Cd in the soil (Fernández-Nieto & Betancourt-González, 2018Fernández-Nieto A, Betancourt-González AR, 2018. Destino sostenible de los residuos generados en las plantas de beneficio avícola. AiBi Revista de investigación, administración e ingenieria 6(1): 11-22. https://doi.org/10.15649/2346030X.473). The SOM at the levels found could have contributed to the concentration of total N in the soil maintaining high values (SEMARNAT, 2002SEMARNAT, 2002. Official Mexican Standard NOM-021-RECNAT-2000 that establishes the specifications of fertility, salinity and classification of soils, study, sampling and analysis. Secretary of Environment and Natural Resources, Mexico, 85 pp.), which is an important element in the inorganic nutrition of plants and improving product quality.
On the other hand, the ECEC (19.9 cmol(+) kg-1) is considered high (Cabalceta & Molina, 2006Cabalceta G, Molina E, 2006. Niveles críticos de nutrimentos en suelos de Costa Rica utilizando la solución extractora Mehlich 3. Agronomía Costarricense 30(2): 31-44.), while the exchangeable K is within the critical (optimal) level and the exchangeable cations Ca and Mg are sufficient in a soil classified as Inceptisol (Cabalceta & Molina, 2006Cabalceta G, Molina E, 2006. Niveles críticos de nutrimentos en suelos de Costa Rica utilizando la solución extractora Mehlich 3. Agronomía Costarricense 30(2): 31-44.). The microelements (Cu, Zn, Fe and Mn) presented high concentrations exceeding the critical levels described for each of them by Calbacete & Molina (2006)Cabalceta G, Molina E, 2006. Niveles críticos de nutrimentos en suelos de Costa Rica utilizando la solución extractora Mehlich 3. Agronomía Costarricense 30(2): 31-44..
The Zn content in the soil can interfere with the adsorption and absorption of Cd due to its chemical similarity (Sikora & Wolt, 1986Sikora FJ, Wolt J, 1986. Effect of cadmium and zinc treated sludge on yield and cadmium-zinc uptake of corn. J Environ Qual 15: 340-345. https://doi.org/10.2134/jeq1986.00472425001500040005x), since they are competing elements and therefore the Zn content negatively affects the uptake of bioavailable Cd by the plant. It could be verified in this work (Table 6a), where Mallows’ Cp demonstrated that Zn is among the variables that predict the bioavailability of soil Cd (Huamaní-Yapanqui et al., 2012Huamaní-Yupanqui HA, Mansilla-Minaya LG, Florida-Rofner N, Neira-Trujillo GM, 2012. Presencia de metales pesados en cultivo de cacao (Theobroma cacao L.) orgánico. Acta Agronómica 61(4): 339-344.; Alloway, 2013Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2).
The plots evaluated showed sandy loam, loam and sandy clay loam textures (Soil Survey Staff, 2014Soil Survey Staff, 2014. Keys to Soil Taxonomy, 12th Edition, USDA-Natural Resources Conservation Service, Washington DC.). According to the granulometry found of sand, silt and clay (Table 2) the study did not present textures with a high percentage of clay, but abundant in sand, similar to those found in studies carried out in Ecuador by Moreno et al. (2018)Moreno J, Sevillano G, Valverde O, Loayza V, Haro R, Zambrano J, 2018. Soil from the coastal plane. In: The soils of Ecuador, pp: 27-77. Springer, Cham. https://doi.org/10.1007/978-3-319-25319-0_2 representing a very unfavorable factor because there may be a high mobilization of Cd moving by leaching to underlying surfaces at the depth of 30 cm (Kruszewski et al., 2018Kruszewski B, Wiesław Obiedziński M, Kowalska J, 2018. Nickel, cadmium and lead levels in raw cocoa and processed chocolate mass materials from three different manufacturers. J Food Compos Anal 66: 127-135. https://doi.org/10.1016/j.jfca.2017.12.012).
A high level of tolerance was observed for Cdleaves (Table 3), according to Kabata-Pendias (2010)Kabata-Pendias A, 2010. Trace elements in soils and plants (4th ed). CRC PRESS, 548 pp. https://doi.org/10.1201/b10158 and Alloway (2013)Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2, who recommend a concentration between 0.05 and 0.5 mg kg-1. According to Barraza et al. (2017)Barraza F, Schreck E, Lévêque T, Uzu G, López F, Ruales J, et al., 2017. Cadmium bioaccumulation and gastric bioaccessibility in cacao: A field study in areas impacted by oil activities in Ecuador. Environ Pollut 229: 950-963. https://doi.org/10.1016/j.envpol.2017.07.080 and Arguello et al. (2019)Argüello D, Chavez E, Lauryssen F, Vanderschueren R, Smolders E, Montalvo D, 2019. Soil properties and agronomic factors affecting cadmium concentrations in cacao beans: A nationwide survey in Ecuador. Sci Total Environ 649: 120-127. https://doi.org/10.1016/j.scitotenv.2018.08.292, the Cd concentration values in leaves can be used as an approximate diagnosis, when the cocoa crop does not present its fruits. This may mean that there is a 3:1 ratio (cocoa leaves: beans).
A statistical correlation was found between the total Cd in the soil and the Cdgrains (Table 4). Ramtahal et al. (2016)Ramtahal G, Yen IC, Bekele I, Bekele F, Wilson L, Maharaj K, et al., 2016. Relationships between cadmium in tissues of cacao trees and soils in plantations of Trinidad and Tobago. Food Nutr Sci 07(01): 37-43. https://doi.org/10.4236/fns.2016.71005 reported similar results on cocoa plantations in Trinidad and Tobago. The results indicate that we can predict the amount of Cd in fruits and grains based on the amount of bioavailable Cd.
Cdleaves did not show a significant correlation with Cdbd in the soil, but when combined with other variables, it was significant (Table 6b). This may be because other metals, such as Al, can interact with the adsorption of this metal by the plant (Cameselle & Gouveia, 2019Cameselle C, Gouveia S, 2019. Phytoremediation of mixed contaminated soil enhanced with electric current. J Hazard Mater 361: 95-102. https://doi.org/10.1016/j.jhazmat.2018.08.062). Other authors, such as Huaraca-Fernández et al. (2020)Huaraca-Fernández J, Pérez-Sosa L, Bustinza-Cabala L, Pampa-Quispe N, 2020. Organic amendments on the immobilization of cadmium in contaminated agricultural soils: a review. Technol Inform 31(4): 139-152. https://doi.org/10.4067/S0718-07642020000400139 refer to the fact that Cdbd in soil is associated with ions (e.g., carboxyls), and these are related to SOM, which by increasing its contents decreases the concentration of bioavailable Cd and affects Cdleaves (Table 6b).
It is worth mentioning that according to the European Union there are still no tolerance levels for Cd in cocoa beans (Barraza et al., 2017Barraza F, Schreck E, Lévêque T, Uzu G, López F, Ruales J, et al., 2017. Cadmium bioaccumulation and gastric bioaccessibility in cacao: A field study in areas impacted by oil activities in Ecuador. Environ Pollut 229: 950-963. https://doi.org/10.1016/j.envpol.2017.07.080; Furcal-Beriguete & Torres-Morales, 2020Furcal-Beriguete P, Torres-Morales JL, 2020. Determinación de concentraciones de cadmio en plantaciones de Theobroma cacao L. en Costa Rica. Revista Tecnología En Marcha 33(1): 122-137. https://doi.org/10.18845/tm.v33i1.5027) so the Cd level is taken as a reference for chocolate with total dry matter content ≥ 50% of the Codex Alimentarius. This should not exceed 0.80 mg kg-1 (FAO/WHO, 2017). According to the European Union in its regulation No. 488 (EC, 2014EC, 2014. Regulation (EC) No 488/2014 amending Regulation (EC) No 1881/2006 as regards the maximum level for cadmium in foodstuffs. European Community Regulation, Belgium, 138/78.) this level may also apply to cocoa fruit. In this study the Cdgrain was well below this limit, with an average of 0.25 mg kg-1.
Similarly, it could be found in Table 2 that the concentration of total and bioavailable Cd in the soil could be related to the average concentration of Cd found in the leaves, fruits and cocoa beans (Table 3), since The Cd transfer coefficient of the soil and plant is 1.0-10.0 (Alloway, 2013Alloway BJ, 2013. Sources of heavy metals and metalloids in soils. In: Heavy metals in soil; Alloway BJ (Ed.), pp. 11-50. Dordrecht: Springer. https://doi.org/10.1007/978-94-007-4470-7_2).
Conclusions
⌅The concentrations of bioavailable cadmium are within the levels of precision and accuracy of the analytical equipment used, for the 21 Inceptisol soil plots extracted at a depth of 30 cm and are relatively low despite the fact that the average pH level is very acid.
The content of soil organic matter presented medium levels, favoring the non-solubility of interchangeable Al and bioavailable Cd.
The models developed for bioavailable Cd, Cd in cocoa leaves, fruits and beans can be used to correct some chemical and biological properties of the soil that can be managed at the farm level, including liming to increase pH and adding organic material to increase the SOM content.