Amonification of by-products from African oil palm fruits (Elaeis guineensis, Jacq.)

Nataly Castro-Ucross¹, Fabio Garrido-Ricoveri¹, Rover Maldonado-Cruz², Omar Araujo-Febres¹, Juan Vergara-López¹ and Rodrigo Martínez Jaramillo¹

¹ Departamento de Zootecnia, Facultad de Agronomía, Universidad del Zulia, Maracaibo, Zulia
juanvergaralopez@gmail.com
² Estación Local El Guayabo, Instituto Nacional de Investigaciones Agrícolas, Catatumbo, Zulia. Venezuela

Abstract

To determine the effect of dry ammonification on the chemical composition and in vitro digestibility of the fruit mesocarp (PFM) and bunch rachis (PBR) of the African oil palm, an experiment was conducted at the National Institute of Agricultural Research (INIA), El Guayabo Local Station, in the municipality of Catatumbo, Zulia State, Venezuela. The byproducts were dry-ammonified with five levels of urea (0, 15, 30, 45 and 60 g.kg DM^-1) over two incubation periods (14 and 28 days) in a completely randomized experimental design under a 2 × 2 × 5 factorial arrangement with 3 replicates. Dry matter (DM), crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF), lignin (Lig) and in vitro dry matter digestibility (IVDMD) were determined. The results show that DM decreased (p<0.01) as the urea dose increased. The lowest values of NDF, ADF, and Lig were observed in the PBR. CP increased (p<0.01) with increasing urea doses, being higher (p<0.01) in the PFM at 28 days of incubation. The highest IVDMD was observed in PBR compared to PFM and was similarly affected by urea and incubation time. Dry ammonification positively altered the chemical composition and digestibility of the byproducts, with PBR being the most favorably affected and thus showing greater potential for use in ruminant feed.

Keywords: animal nutrition, cluster, chemical composition, food science, mesocarp

Introduction

The African oil palm (Elaeis guineensis Jacq.) is a perennial crop, well-suited to the soil and climate conditions of the tropics. The processing of the fruits of this plant species generates harvest and agro-industrial by-products, such as the fruit mesocarp (PFM) and the bunch rachis (PBR) which, being lignocellulosic materials, have potential for use as ruminant feed.

The vast majority of fibrous agricultural residues are characterised by being available in varying quantities, with high proportions of fibrous fractions that are slow to degrade and have low protein content, which negatively affects their digestibility and voluntary intake by the animal (Cuesta et al 2000); therefore, before they can be effectively used in feed rations, they require further processing (Zahari and Alimon, 2005).

One option that would improve the nutritional value of PFM and PBR is the ammoniation; this process adds nitrogen in the form of ammonia to these materials, resulting in an increase in crude protein content and improved digestibility. Furthermore, ammonia breaks down lignocellulosic bonds, leading to the release of hemicellulose, which improves digestibility. This treatment, in addition to being one of the most economical, is easy to carry out and has already yielded known results for other plant-based materials (Cuesta et al 2000; Ventura et al 2002). The aim of this study was to evaluate the effect of dry ammoniation on the chemical composition and in vitro digestibility of two by-products from the processing of African oil palm fruit.

Materials and methods

The experiment was carried out at the El Guayabo Local Station of the Instituto Nacional de Investigaciones Agrícolas (INIA), located at the south of Lake of Maracaibo, 5 km from the Machiques-Colón junction and 9 km from El Guayabo (geographically at 08°39’16 ’ north latitude and 72°19’56’’ west longitude), in the municipality of Catatumbo, Zulia state (Venezuela). The area is classified as tropical rainforest, with an annual rainfall of 1802.5 mm·year⁻¹, an average annual temperature of 28 °C and relative humidity of 80%.

The PFM and PBR samples (80 kg of each material) were obtained from the extraction line at the Palmeras Diana del Lago Industrial Complex, located 35 km from Casigua el Cubo, in the Jesús María Semprún municipality of Zulia State, Venezuela. The material was dried for 10 days in a room at 65 °C with forced ventilation. The PBR was chopped until it reached a size similar to that of the PFM particles. To ammoniate the fibrous materials, 60 plastic containers with a capacity of 18 L were used. Galvanised wires were attached to the inside of the plastic lids of the containers, which were subsequently used to tie the mesh bags containing the by-products to be ammoniated. Samples of 200.00 g of dried and chopped PFM and PBR were weighed and placed into bags made of nylon mesh, which were tied at the ends to prevent material loss. The bags were suspended using the galvanised wire attached to the container lid.

Treatments

Two by-products (PBR and PFM), five urea levels (0, 15, 30, 45 and 60 g urea·kg DM⁻¹) and two ammonification periods (14 and 28 days) were evaluated, resulting in a total of 20 treatments.

Urea solutions corresponding to the five treatments levels (0, 15, 30, 45, 60 g urea·kg DM^-1) were prepared. To maintain a rate of 420 mL·kg DM⁻¹ (equivalent to 84 mL per container), the solutions were formulated to achieve concentrations of 0, 3.57, 7.14, 10.70 and 14.29 % w/v. The corresponding solution and 5.0 g of fresh, cut grass (Brachiaria arrecta), which acted as a ureolytic agent, were placed at the bottom of each container. The lids were fitted and the containers sealed airtight, then stored in the shade in a covered shed for 14 and 28 days, respectively.

Once the ammonification periods had elapsed, the bundles were removed from the containers, left to air for 12 hours and stored in appropriately labelled paper bags with perforations to allow drying in a forced-air oven at 65°C for 48 hours. Once dry, the samples were ground in a Willey mill with a 1 mm sieve and stored in labelled, airtight plastic bags.

The dried and ground samples were subjected to laboratory analysis to determine dry matter (DM), crude protein (CP), neutral detergent fibre (NDF), acid detergent fibre (ADF) and lignin (LIG) (AOAC, 1990a and b).

In vitro digestibility was determined using F57 nylon filter bags (ANKOM Technology), into which 0.25 ± 0.005 g of each material was weighed directly into the bag and heat-sealed. In vitro digestibility was determined using the Tilley & Terry (1963) method, modified by Minson & McLeod (1972), using an Ankom Daisy II® incubator.

Statistical design and data analysis

The experimental design was completely randomised in a 2×5 factorial arrangement, with three replicates. The factors evaluated were ammonification time, type of material to be ammonified and urea levels; whilst the dependent variables were the contents of DM (%), CP (%), NDF (%), ADF (%), LIG (%) and IVDMD (%). The data were recorded in MS-Excel^® spreadsheets. The data were subjected to analysis of variance using the General Linear Model, employing Infostat software (Infostat, 2008). Those variables that proved significant (p<0.05) were subjected to Tukey’s test with probabilities of up to 5% (Infostat, 2008).

Results and discussion

Dry matter (DM)

Analysis of variance revealed that DM content was affected by the interaction between urea dose × by-product × incubation time. As the urea dose and incubation time increased, DM decreased (p<0.05) in both by-products. However, the control treatments increased their DM content. This behaviour is attributed to the alkaline effect of ammonium on the hydrolysis and partial solubilisation of cellulose, as well as the breaking of lignin-hemicellulose ester bonds (Rodríguez et al 2002). This reaction results in a double loss of mass: the volatilisation of residual gaseous compounds and the solubilisation of carbohydrates that are washed away or lost as effluents during the drying process, leading to a net decrease in DM (Ruiloba and Solís 2022).

The PBR yielded the highest DM levels compared to the PFM. Both results are higher than the 86.2% DM reported by Zahari and Alimon (2005) for compressed palm fibre (mesocarp). The DM contents were higher after 28 days of ammoniation than after 14 days, a result similar to that reported by Camacho et al (2003) for Brachiaria humidicola hay ammoniated for 21 days, at 92.4%.

The main effect of urea resulted in a consistent decrease in DM from 92.3% for 0 g urea·kg DM⁻¹ to 90.2% for 60 g urea·kg DM⁻¹. This pattern is consistent with that reported by Noersidiq et al (2020) in ammonium-treated African palm trunks. In contrast, Ruiloba and Solís (2022) found increases in DM in fresh sweet potato tubers as the urea concentration was increased from 1.50% (25.1% DM) to 4.50% (26.0% DM), suggesting a difference in the reactivity of ammonium towards the fibrous matrix of the palm by-product compared to ammonia.

Crude protein (CP)

The analysis of variance did not detect any differences in the three-way interaction (p > 0.05); however, increases in CP content were observed as urea doses were increased, regardless of incubation times (Table 2). This is consistent with the results of Hurtado et al (2021), who, when evaluating the effects of ammonification on sugarcane bagasse at inclusion levels of 0, 3 and 5% at two fermentation times (30 and 44 days), found increases in CP content as inclusion levels rose from 3.78% to 29.3%. They also found that, at 44 days, CP content was lower, likely attributable to the microorganisms’ own nitrogen requirements for growth. Meanwhile, Castellanos et al (2017), evaluating the effect of three levels of urea (0, 3 and 6%) on maize husks (fibrous residues), found increases in CP content from 5.08% to 12.9%, consistent with this study.

PFM yielded the highest CP levels regardless of the urea concentrations applied, with incremental increases in this variable observed in both cases as the urea dose was increased (Table 3). Similarly, PFM yielded the highest CP results, irrespective of the incubation times applied (Table 4), although longer incubation times favoured CP concentration in both by-products. Similar results were obtained by Ponce and Romero (2015), who, using a dosage of 3 kg of urea in 20 L of water to ammoniate maize stalks, recorded 12.6% CP at 21 days, which subsequently increased to 14.2% at 28 days.

Ventura et al (2002) found increases in CP content in sorghum stubble hay when subjected to dry ammoniation with doses of 20, 40 and 60 g urea·kg ^-1 of hay (9.29% on average) compared with the control (5.22%), explaining that the moisture content of the treated ammoniated material may favour the action of the alkali on the cell walls of the forage. However, it should be noted that urea is a source of non-protein nitrogen, which would be utilized for protein synthesis by rumen microorganisms and degraded by proteolytic enzymes to form peptides and amino acids (Noersidiq et al 2020).

Neutral Detergent Fiber (NDF)

Analysis of variance revealed that the NDF content was influenced by the interaction between by-product dose × urea × incubation time, showing fluctuating behaviour between each dose (Table 1). NDF reached its lowest numerical value (32.9%) with the application of 60 g·kg DM⁻¹ of urea at 28 days in the PBR, representing an absolute reduction of 6.12 percentage points compared to the 14-day incubation period (39.0%), with no difference in means between them (p>0.05). On the other hand, the PFM obtained the highest NDF contents (74.1%) when 60 g·kg DM⁻¹ of urea was applied, showing no difference from the other urea applications.

In the PFM and PBR under both incubation times, a fluctuating trend persists, amplified by the anatomical heterogeneity of the by-products. This difference in their composition causes the effect of NH ₃ to be differentially reactive. Therefore, the fluctuations observed in the data, particularly at high doses, reflect the combination of chemical saturation of ammonia and variability in the proportion of more lignified tissues that are less reactive. This is consistent with Cuervo and Gutiérrez (2018), who observed no differences between ammoniated maize and ensiled and ammoniated maize, with an average of 72.3% between the two.

Regarding the urea × incubation time interaction, differences were observed only between the application of 30 g·kg DM⁻¹ at 14 days (60.4%) and that of 60 g·kg DM⁻¹ at 28 days (51.1%). The rest did not differ from one another, averaging 56.8%. This is confirmed by the fact that the simple effect of incubation time did not produce differences (p>0.05) in NDF values.

Valbuena (2006), when evaluating different urea concentrations (3, 5 and 7%) in the ammoniation of Paspalum fascilutatum, determined a trend toward a decrease in crude fiber as the urea concentration increased, averaging 30.6%; whereas the results of Rodríguez et al (2002) for B. humidicola cut at different ages and ammoniated with 0% and 6% urea—78.5% and 75.5%, respectively—contrast with the downward trend in NDF observed in relation to increasing urea levels, which was not observed in this study.

It is believed that this variability stems from the fact that, in palm byproducts, the high density and lignocellulosic nature of the matrix create a physical barrier that restricts the diffusion of NH ₃ into the interior of the material, limiting its effect to the surface layers. In contrast, the porous structure of tropical grasses could facilitate a more homogeneous penetration of ammonia, explaining the differences observed in both substrates. PFM had the highest NDF content (70.6%) in contrast to PBR (42.5%). This NDF value for PFM is reasonable, as it is a fraction of the palm fruit with a high content of cellulosic fibers responsible for oil synthesis and retention.

It is important to note that the efficiency of ammonification depends on the physical preparation of the sample. Although the PBR was ground to achieve the particle size of the PFM, the final microstructure may differ due to its anatomical composition. In this regard, Tian et al (2023) demonstrated that different milling methods in cereals alter particle size and porosity, impacting the enzymatic accessibility of starches and digestibility. In this context, it is hypothesized that PBR, being more fibrous, maintains a porosity that facilitates greater NH₃ diffusion. In contrast, it is hypothesized that PFM, with a potential content of soluble components in the form of residual oils, could form a more compact and impermeable matrix after grinding, contributing to the morphological variability of the byproducts and would partly explain the limited response of urea on NDF.

Acid Detergent Fiber (ADF)

The by-product × urea × incubation time interaction affected ADF values (Table 1), exhibiting the same inconsistent behavior as NDF. The lowest ADF contents were obtained in the PBR, regardless of the dose and incubation times applied; under incubation times of 28 days, ADF contents decreased (p<0.05) as the urea dose was increased from 0 to 60 g urea·kg⁻¹ DM. The ADF contents contrast with those obtained by Segura et al (2001) at 54.2% and resemble those of Vergara and Araujo (2006) in B. humidicola cut during the dry season (37.9%).

The ADF values reported by Conde et al (2000) in PFM treated with 5, 10, 11, and 22% urea and (NH₄)₂SO₄ for 30 days are higher than those obtained in PFM treated with urea in this study; however, it should be noted that the value of (NH₄)₂SO₄ at 22% (50.8%) is similar to that of 60 g urea·kg⁻¹ DM at 28 days (51.9%). These authors used the same sources and doses, but by ammoniating the solids retained in the primary and secondary screens of the palm oil industry process, they obtained results similar to those obtained in the empty bunch at 14 and 28 days.

These results can be explained, on the one hand, by the relative decrease in fermentable carbohydrates, including soluble carbohydrates, during the incubation period (Fernandez et al 2021) and, on the other hand, by the limited transfer of gaseous NH₃ corresponding to the byproducts used, preventing a linear response to increased urea doses, especially in the PFM, which has a denser matrix.

Lignin (LIG)

The analysis of variance did not detect any significant effects (p> 0.05) regarding the by-product × urea × incubation time interaction.

Regarding the interaction between incubation time and urea, it was observed that at 14 days, incremental doses of urea tended to decrease LIG content to a maximum of 14.3% when 45 g·kg DM^-1 of urea was applied. In contrast, at 28 days, a decrease was observed from 0 g·kg DM⁻¹ (15.9%) to 60 g·kg DM⁻¹ (14.0%) (Table 2).

These data are similar to those collected by Madrid et al (1998) in untreated pumpkin byproducts (13.3%) and could be explained by the limitation imposed by LIG’s natural resistance to hydrolysis. Similar responses to the effects of incubation time and urea levels have been reported in sugarcane bagasse, where LIG decreased from 7.74% (control) to 4.75% upon application of 5% urea (Hurtado et al 2021) and B. humidicola , when treated with 0, 3 and 6% urea for 28 days, showed decreasing LIG values of 5.23, 4.64 and 4.49%, respectively (Rodríguez et al 2002).

The types of byproducts resulted in differences in LIG content. PBR yielded a material with low LIG content (8.30%) compared to PFM (22.1%) and this is lower than the 20.0% LIG reported in PBR used for compost by Segura et al (2001), perhaps due to greater efficiency in oil extraction. The LIG value in PBR is similar to the values obtained in ammoniated B. humidicola (9.5% LIG) (Atencio et al 2008) and in endosperm meal produced by mechanical extraction of palm fruit oil (8.6%) (Vargas and Zumbado, 2003).

Despite the statistical significance observed in the interactions, the low magnitude of the LIG content is an expected result. LIG not only resists hydrolytic action (Chávez-Sifontes and Dómine, 2013), but also acts as a physical barrier that can restrict the diffusion of gaseous NH₃ toward cellulose and hemicellulose. Therefore, this behavior, although significant, is limited by the density of the lignocellulosic fraction of the PFM, reaffirming that the limitation on gaseous NH₃ transfer and the heterogeneity of the byproduct are the factors that most strongly influence the efficiency of ammonification.

In vitro dry matter digestibility (IVDMD)

Analysis of variance revealed highly significant effects of the byproduct × urea × incubation time interaction. The highest IVDMD was observed when 15 g of urea was applied to PFM with a 28-day incubation period (72.7%). However, it should be noted that the lowest IVDMD were obtained in PFM with 30, 45, and 60 g of urea at 14 days and with 30, 60 and 45 g of urea at 28 days (Table 1). Similar results were obtained by Camacho et al (2003), who reported a 7 percentage-point increase in the in vivo digestibility of DM in Brachiaria humidicola hay —both untreated and treated with 10% urea— after 21 days of incubation.

This inconsistency may be due, on the one hand, to the denser matrix of PFM, with a potential concentration of residual oils that impede the diffusion of NH₃ in the matrix and consequently, resist the hydrolytic action of urea, as well as to the in vitro system used, in which increasing levels of urea and incubation time for decomposition raise the ammonia soluble in the medium’s moisture, thereby inhibiting microbial growth due to higher ammonia concentrations and increased pH levels, which are unfavorable for the development of the ruminal microbiota of the inoculum (González et al 2012).

Regarding the byproduct × incubation time interaction, the IVDMD was higher in the PBR (Table 4). Likewise, regarding the urea × byproduct interaction, it was observed that the IVDMD of the PFM without urea application was higher than in the urea applications, with the exception of the 15 g dose, and that the remaining applications (30, 45 and 60 g of urea) were lower than those obtained in the PBR (Table 3), these increases in the PBR are attributed to a greater effect and exposure of ammonium on the cell walls and are confirmed in the byproduct × incubation time interaction, where the PBR obtained an average of 61.65% at both time points, while the PFM obtained 54.6% at 28 days and 45.4% at 14 days of incubation.

Regarding the simple effect of the byproduct, PBR presented the highest IVDMD value (61.7%) in contrast to PFM (50.0%), with the PBR value differing from that reported by Segura et al (2001), who found 35% IVDMD in the same material However, Zahari et al (1999) report digestibility values of 60 to 62%, very similar to those reported in this study. The IVDMD and LIG values obtained from the empty cluster demonstrate the promising potential of this byproduct in ruminant feed.

Rodríguez et al (2004) observed increases in IVDMD from 65.1% to 70.8% in B. humidicola treated with 0% and 6% urea. When ammonification occurs satisfactorily, the crystalline structure of cellulose breaks down, as do the bonds between it and the hemicellulose-lignin fraction, releasing a greater proportion of soluble carbohydrates, proteins and minerals while simultaneously increasing the size and surface area of structural carbohydrates exposed to attack by ruminal microflora (Fuentes et al 2001; Arce et al 2003; Camacho et al 2003; Rodríguez et al 2004; Barrios and Ventura 2005).

Conclusions

Dry ammonification treatments applied to PFM and PBR reduced the dry matter (DM) and lignin (LIG) contents and increased the crude protein (CP) content.

Although PFM is superior from a nutritional standpoint, the reduction in NDF and FDA contents was limited and no clear linear response to increased urea dosage was observed, indicating the need to optimize treatments for these lignocellulosic products.

Increasing the urea concentration in the dry ammonification treatment significantly increased the in vitro dry matter digestibility (IVDMD) of the ammonified PBR and PFM.

The IVDMD of PBR was higher than that of PFM; therefore, it shows greater potential for use in ruminant feed.

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