The effects of feed supplemention on the in vitro fermentability, ration digestibility and methane gas production across different dairy cattle feeding rations at the farms level

Zahra W A¹, Suharti S², Jayanegara A², Akbar I³, Armandani S³, Wardani W W³ and Permana I G²

¹ Department of Animal Production Technology, Faculty of Animal Science, IPB University, Bogor 16680, Indonesia
permana@apps.ipb.ac.id
² Department of Animal Nutrition and Feed Technology, Faculty of Animal Science, IPB University, Bogor 16680, Indonesia
³ Nutricell R&D, Cibis Nine 12^th Floor Unit G1, Jl. TB Simatupang No. 2, Jakarta 12560, Indonesia

Abstract

The escalating demand for livestock products must be accompanied by sustainable livestock management systems to minimize greenhouse gas (GHG) emissions. Adding feed supplement to subsequently increase feed quality and efficiency emerges as a viable strategy in mitigating GHG emissions. The study aimed to evaluate the effects of feed supplementation at different dairy cattle feeding rations on the in vitro fermentability, ration digestibility, and methane gas production. The research was conducted in October-December 2022 in three dairy cattle farms in East Java. The nutrient composition of the raw materials being commonly used at the farms and the effects of feed supplement on dairy cattle rations were analysed. An experimental design was conducted using a Randomized Block Design. The treatments included the use of feed supplement (P1) and without feed supplement(P0). Three different rations which were including KTSPSR, KTRA, and KUDS and four replications at each ratio were used. The dry matter (DM), crude protein analysis (CP), crude fibre (CF), in vitro procedures including volatile fatty acids (VFA) analysis, CH₄ gas production and NH₃were analysed. The results showed that the feed content of KTRA has the lowest CP and the highest CF content. However, the KUDS has the highest CP and the lowest CF content. The effects of feed supplementation showed a normal rumen pH and significantly affected (p<0.05) the concentration of N-NH₃ in the rumen, the total VFA concentration, the dry matter digestibility (DMD) and organic matter digestibility (OMD). To conclude the use of feed supplement significantly reduced pH, increased N-NH₃ concentration, total VFA, and ration digestibility.

Keywords: feed supplement, feed efficiency, methane emissions

Introduction

Livestock sector plays an essential role in food sovereignty. The escalating demand for livestock products, especially milk, demonstrates an annual increase attributed to population growth, the expanding middle to upper-class demographic, and heightened awareness concerning the animal protein consumption. However, the increasing demand for milk consumption is not followed by the national milk production supply. Recently, the dairy cow population in Indonesia stands at approximately 650,000 heads, with national milk production of 1.5 million tons per annum but the aggregate national milk output merely fulfils 18% of the total milk demand (Indonesian Central Statistics, 2021).

Dairy farming is concentrated within Java Island, with West Java, Central Java, and East Java provinces emerging as the milk production centre. East Java has the highest dairy population and milk production, compared to the other two provinces, counting a substantial amount of 302,300 cows and an annual milk production of 558,758 tons (Indonesian Central Statistics, 2021). The domination of dairy farming in Java island leads to two critical issues, i.e., manure management and availability of feeding due to land limitations for livestock production (Al Zahra et al 2020).

Feed quality, both forage and concentrate is vital to improve milk production (Johansen et al 2018). On average, the dairy farmers have 0.44 hectares land for dairy farming and it fulfils only 63% of the requisite feed demand. To fill the gap, the farmers must find roadside grass, and particularly in the dry season, when there is often a lack of forage supply, leading to an acute deficiency in forage supply. Consequently, the farmers must buy forage from other areas. Concentrate, as another primary feed source for dairy cattle and often it varies between farms in terms of quality.

Improving feed quality is necessary to improve the performance of dairy cattle and this is also closely related to reducing greenhouse gas (GHG) emissions in the dairy cattle supply chain (Waghorn et al 2011; Gerber et al 2013; Hawkins et al 2021). Globally, the livestock sector contributes 14.5% of GHG emissions from human anthropogenic activities. GHG emissions analysed by using Life Cycle Assessment (LCA) estimates that feed is the main contributor (47%) to total dairy cattle production chain emissions due to its association with CH ₄ emissions. Waste management is another important factor (26%), contributing to the total emissions in the dairy cattle production chain due to its association with CH₄ and N₂O emissions. A study by Apdini et al (2021) estimated that total emissions in Indonesia's dairy cattle supply chain from the enteric fermentation reached 55-60%, with waste management accounted for 12-15%.

Improving feed quality can be done, one of which, through feed supplementation. Feed supplement plays an essential role in the physiological process of livestock nutrition related to health, growth, reproduction, and hormone systems (Soetan et al 2010). The effectiveness and practical application of feed supplement faced limitations due to the dependency on the diet's response. Factors such as the availability of feed supplement in the basal ingredients significantly influenced the performance and methane reduction (Weiss, 2017). These complexities made it challenging to achieve consistent and predictable feed supplementation results for dairy cows.

Improving feed quality using feed supplement is considered a mitigation strategy to reduce GHG emissions from dairy farming (Feng et al 2020; Hristov et al 2013). Research on the effects of feed supplementation on feeding patterns which especially reflecting the condition at the farms level has yet to be extensively conducted. Whereas this information is essential to design mitigation strategies to reduce emissions from smallholder dairy farms which subsequently contribute to sustainable dairy production in Indonesia. Hence, this study aims to evaluate the effects of feed supplement on the in vitro fermentability, ration digestibility, and methane gas production across different dairy cattle feeding rations at the farms level.

Methods

Data collection

The research was conducted from October to December 2022 at dairy farms in East Java. This study was executed in two distinct stages. In the first stage, the raw materials of concentrate, commonly used in dairy farms in East Java, were analysed to understand the nutrient content. Subsequently, in the second stage, the effect of feed supplement was analysed, allowing for a comprehensive evaluation of the in vitro fermentability, ration digestibility and CH₄ gas production.

First stage

The concentrate feed ingredients commonly used by dairy farmers in East Java were collected. These raw materials were procured from the largest dairy cooperative that serves as a primary supplier of concentrate feed within the East Java province. The collected materials include wheat bran, wheat pollard, rice bran, corn gluten feed, distiller's dried grains with soluble (DDGS) corn, palm kernel meal, palm kernel expeller (PKE) pellet, copra meal, copra pellet, and coffee hulls Nutrient analysis of the ingredients including the dry matter (DM), ash, crude lipid (CL), crude protein (CP), crude fibre (CF), and nitrogen-free extract (NFE) were analysed.

Second stage

A comprehensive investigation was undertaken through a survey and in-depth interviews involving three dairy farms in Malang, Kediri, and Blitar districts. These particular districts were chosen due to the high dairy cattle population compared to other districts in East Java province. The included farms group in this study were: Sumber Rezeki Dairy Cattle Farmer Group in Malang (KTSPSR), Risin Agung Livestock Group in Kertajaya-Kediri (KTRA) and Semen Village Cooperative Unit in Blitar (KUDS) (Figure 1). We selected a farmer leader from each respective farm group. KTSPSR located in Batu, Malang, and has been established since 2016. KTRA situated in Kediri and has 14 livestock group members. KUDS, located in Blitar, has 649 active members and has been operational since 1981 but began raising cattle in 2010. The cattle population in KUDS is 2,774 cows, in KTSPSR is 850 cows and in KTRA is 1,500 cows.

We identified respondent profile which included their dairy farming experience, educational background, and occupational status. Following this, the type of feed and feeding management for each farm was observed by direct observation. Information about feed provision (kg animal^-1day^‑1) and milk production (kg animal^-1day^-1) for each lactating cow, and manure management systems, was obtained through interviews with the farmers. In order to facilitate nutrient analysis, approximately 500 grams of feed and fresh manure samples were collected from each farm.

Experimental diet of feed supplement

An experimental design was carried out to investigate the effects of feed supplement on in vitro fermentability, ration digestibility, and CH₄ gas production. In an vitro trial, the experimental design used a randomized block design of 2 x 3 x 4 factorial consisting of two supplements, three feed ration types, and four replications. Two supplementations consisted of the use of feed supplement (P1) and without feed supplement (P0). Feed supplement, i.e., Rumnesia Dairy from PT Nutricell Pacific was introduced at the level of 15% of the total diets at each feeding ration and this feed supplement has been used by these farmers. Three types of feed ration included the KTSPSR, KTRA, and KUDS, and four replications used in this experimental diet.

Laboratory analysis

The laboratory analysis was conducted to analyze the nutrient content of the treatments, which consisted of moisture, crude protein, and fat content. In order to analyze feed fermentability and digestibility, the in vitro procedure, as described by Tilley and Terry (1963), was used. The in vitro analysis included the VFA analysis, CH₄ gas production, and N-NH₃ content. The in vitro procedure was carried out to analyse feed fermentability and digestibility. This procedure was performed using the in vitro method according to the Tilley and Terry (1963) method. For the fermentability analysis, 0.5 grams of the sample were combined with 40 ml of buffer McDougall liquid and 10 ml of rumen liquid. The mixture was then infused with CO₂ gas and incubated at 39^oC. After a four-hour incubation period, HgCl₂ was introduced, followed by centrifugation at a speed of 3000 rpm for 15 minutes. The resulting supernatant was tested to determine the rumen pH, Volatile Fatty Acid (VFA) levels, and N-NH₃. Subsequently, CH₄ gas was analyzed using gas chromatography. In the digestibility analysis, the experimental diets were incubated in rumen liquor for 48 hours to simulate the fermentation process that occurs in the rumen. Then, the samples underwent digestion with an HCl-pepsin enzyme solution to imitate the enzymatic process. After digestion, the dry and organic matter residues were subtracted from the samples to calculate the value of in vitro digestibility (dry matter and organic matter).

In addition, manure samples were collected, and the analysis of nitrogen (N) and N-NH₃ content was conducted. The determination of moisture content followed the standard method outlined in AOAC (2005). The standard Kjeldahl method determined the protein and total nitrogen content. The pH meter was used to measure the pH. Total Volatile Fatty Acid (VFA) was quantified using the standard steam distillation technique. Similarly, the concentration of N-NH₃ was evaluated utilizing the micro-diffusion method with a Conway cell (Conway, 1957). The partial CH₄gas production was analysed by using gas chromatography and the total CH₄ gas production quantified using the equation established by Moss et al (2000) (Equation 1)

CH₄ gas production = 0.45C₂ − 0.275C₃ + 0.40C ₄ (Equation 1)

where: CH₄ is methane (mMol), C₂ is acetate, C ₃ is propionate and C₄ is butyrate.

Data analysis

The nutritional composition of dairy concentrate ingredients, respondent profile, feeding management system, milk production, and manure management were presented through descriptive analysis. The effects of feed supplement on pH, N-NH₃, total VFA, VFA partial, the CH ₄ gas production, dry matter digestibility (DMD) and organic matter digestibility (OMD) were analysed using the analysis of variance (ANOVA) conducted through SPSS software (IBM SPSS Statistics, USA). The significance parameters were tested using Duncan's multiple-range test.

Results and discussion

Concentrate feed ingredients analysis

Table 1 presents the nutritional composition of dairy concentrate feed in East Java. The energy sources in the concentrate feed include wheat bran, wheat pollard, and rice bran. The sources of protein in the concentrate feed include corn gluten feed, distiller's dried grains with soluble corn, palm kernel meal, palm kernel extraction pellet copra meal, copra pellet, and coffee hulls. Our results reveals that the CP content of rice bran is relatively lower than that Mila and Sudarma (2021) and Feedipedia (2023). The inclusion of supplementary materials, such as husk fractions, might explain the lower value of rice bran.

Feeding management system

Feeding management in all farms encompassed a combination of forage and concentrate. The availability of forage depends on the seasons, for example in the dry season when the forage is limited from August to October, the farmers have to purchase additional grass at a cost of 400 IDR/kg and corn straw at a rate of 200-800 IDR/kg. Abundant grass was available from December to January. To enhance grass growth, the farmers applied fertilizer, specifically urea, three times annually, with an application rate of 250 kg per hectare on average. This practice aimed to bolster the nutritional content of the grass. The concentrates was procured mainly from the dairy cooperatives and supplemented by additional quantities procured from the commercial sources.

Table 2 shows the feed being provided for dairy cows (in kg as feed) in the three research locations. At KTSPSR, the cows' diet mainly consisted of odot grass, concentrate and legumes. They grew the odot grass on their own land and harvested at an average interval of 90 days. To prepare the grass for feeding, a chopper machine was used to chop it into smaller pieces and a small quantity of legumes, such as lamtoro (Leucaena leucocephala) leaves, as a supplementary feed was added. The cows were fed twice a day – once in the morning and again in the afternoon.

The feed in KTRA was odot and elephant grass, concentrate, and legumes. The forage composition was 80% odot grass and 20% legumes (optional). Kaliandra (Calliandra), Indigofera (Indigofera sp), and lamtoro (Leucaena leucocephala) were commonly used legumes on the farm. The farmer offered the grass three times a day, i.e., morning, afternoon, and evening and this was two times a day for the concentrate, i.e., morning and afternoon.

The feed in KUDS including concentrate and the odot grass, elephant grass, wild grass, and corn straw, depend on the availability of the grass. The grass was cut into smaller pieces using a chopper machine, followed by a fermentation step and subsequently fed to the cows. The feed was offered twice a day—during the morning and afternoon sessions.

Figure 1 illustrates the feed ratio in both as-fed and dry matter in three farms. From Figure 1, KTRA provides the highest feed when considered on an as-fed basis, in comparison to the other farms. However, the provision of feed (as-fed) does not correspond to the quantity of dry matter. At KUD Semen Blitar, despite the relatively lower quantity of feed (as feed) offered to the cows, the farm achieved a greater yield of dry matter.

Figure 1. Feed ratio based on as-fed (a) and dry matter basis (b). The black solid colour shows the total concentrate, and the grey solid colour shows the total forage fed to the lactating cows

Figure 2 shows the proportion of forage and concentrate feed in dry matter basis in KTSPSR, KTRA and KUDS. Based on our study, the proportion of forage and concentrate feed in KTSPSR appears to be balanced. In KTRA and KUDS, the proportion of feed is characterized by a notably higher proportion of forage. The difference in feed proportion upon the diverse feed sources accessible at each respective farm location.

Figure 2. Proportion of forage and concentrate feed in dry matter basis in KTSPSR (a), KTRA (b), and KUDS (c). The black solid colour shows the proportion of concentrate and the grey solid colour shows the proportion of forage

The results of nutrient composition analysis (Table 3) reveal that the feed provided by KTRA has the lowest protein content and the highest crude fibre content. Meanwhile, the feed supplied by KUDS indicates the highest protein but contains the lowest crude fibre content.

Milk production

Table 4 shows the dairy composition characteristics of the observed respondents. The results indicated that the total population was 13 to 32 dairy cattle which is relatively higher compared to the general smallholder farmers with 3 to 5 dairy cattle. The results showed that lactating cows emerge as the predominant category among the dairy types represented in the study.

At the time of the study, the outbreak of Foot and Mouth Disease (FMD) had a negative impact on many dairy farms in East Java, including the dairy farms in this study. This situation led to the unfortunate death of dairy cattle and a subsequent drop in milk production. Consequently, in this investigation, the direct assessment of milk production and feed consumption was avoided to prevent any potential contamination. According to statements from dairy farmers, before the FMD outbreak occurred, the milk production varied between 10 to 16 litres per cow daily. Unfortunately, during the FMD outbreak, there was a notable decrease of 30 to 40% in comparison to the pre-FMD period. Recently, there has been a decline in milk production, with an estimated range of 7 to 10 litres per cow per day (as shown in Figure 3). Under normal circumstances, milk can be supplied to milk processing companies such as PT Nestle Indonesia, PT Indolakto, PT Sari Husada and PT Frisian Flag Indonesia.

Figure 3. Milk production prior to and during the foot and mouth disease outbreak. As the data is collected once during the interview, the standard deviation of milk production is not available

Manure management

In this study, different manure management was observed. In KTSPSR the solid manure (faeces) was composted, yielding an organic fertilizer as the end product. In KTRA and KUDS the biogas technology sized of 12 m³ is used to manage the solid manure in both farms. Across all farms, urine was discharged due to constraints in urine management. The laboratory analysis results of N and N-NH₃ content in manure, compost and bio slurry are shown in Table 5.

The composting process in KTSPSR has increased the N content. This process facilitates a gradual release of N into the soil at a slower rate (approximately 1-3% of total N per year) compared to the application of inorganic fertilizers (Al-Bataina et al 2016). Moreover, biogas technology is becoming more popular due to its dual benefits of efficient dairy manure management and simultaneous reduction of GHG emissions. In this process, organic matter present in the manure is broken down by microorganisms to generate CH₄ gas, which can serve as a fuel source (Meegoda et al 2018). Additionally, bio slurry, a valuable by-product, is produced as an organic fertilizer (Chen et al 2008; Sung and Liu 2003).

The nitrogen (N) content in faeces and urine determines the N₂O formation, and the N in manure is susceptible to loss through processes such as volatilization and N leaching. Manure storage system storage is essential in determining the quality and the N loss. For instance, the open storage of bio slurry in KUDS reduces the N content of bio slurry, while the closed storage system adopted by KTRA prevents N loss. The N-NH₃ content in fresh manure varies among the three observed farms in this study and is likely to be attributed to feed composition, particularly the concentrate, which serves as a primary contributor to N formation (Cai and Akiyama 2016; Rivera and Chará, 2021; Wecking, 2021). The N-NH₃ value in KTRA is the highest among others, which could be explained to the relatively lower level of retained nitrogen (N) within the faeces. Consequently, a more significant proportion of N is susceptible to loss in the form of N-NH₃, leading to the observed higher N-NH₃ content.

Effects of feed supplementation

The effects of diverse feeding rations and feed supplement on DMD and OMD is shown in Table 6. The DMD values of P0 ranged from 562 to 644.6 (g/kg) and this was from 575.4 to 652.3 (g/kg) in P1. The type of diverse feeding rations and feed supplement significantly affected the DMD of the diets (p<0.05) with an observable interaction between the two factors. The KUDS had the highest DMD compared to others feeding rations. The addition of feed supplement has increased the DMD of the KTSPSR by 6.5% (p<0.05) but adding feed supplement did not significantly increased DMD in KTRA and KUDS. The DMD was positively correlated with the energy and protein content of the diets in which the diets with higher energy content tended to exhibit increased DMD values.

The OMD values for P0 ranged from 549.4 to 635.3 (g/kg), and this was 568.2 to 647.6 (g/kg) in P1. The diverse feeding rations and feed supplement significantly affected OMD (p<0.05) with an interaction between these two factors. The KUDS had the highest OMD compared to others feeding rations. Feed supplement significantly increased OMD in Rations KTSPSR by 6.9% and KTRA by 1.8% (p<0.05), but not in Rations KUDS.

The effect of feed supplement on pH, N-NH₃, CH₄ gas production, and total VFA are presented in Table 7. The rumen pH in this study remained within the normal value, ranging from 6.80 to 6.95 and aligned with the study of Akyurek and Salman (2021). Diverse feeding rations and the addition of feed supplement affected pH in the rumen and has decreased the pH value among treatments (p<0.05). The pH is linked to the nutrient content of the diet. The use of feed supplement in this study has decreased rumen pH and this findings exhibit disparities compared to Silva et al (2022), which indicating that the maintenance of rumen pH was achieved through the supplementation of a combination of vitamin A,D, and E and mineral premix. Generally, the decrease in rumen pH is associated with diets containing high concentrate.

The production of N-NH₃ indicates the amount of easily degradable protein in the feed. Diverse feeding rations and the addition of feed supplement affected N-NH₃ in the rumen and has increased the N-NH₃ value among treatments (p<0.05). The inclusion of feed supplement has increased the total VFA concentration in KTSPSR by 31.9% and in KTRA by 11.7%.This and this indicate that the effects of feed supplement vary depending on the feeding patterns utilized. Feeds characterized by higher protein content (KTSPSR and KUDS) yielded correspondingly increased NH₃ concentrations. Feed supplement heightened the fermentability of CP, leading to an increase in the concentration of rumen NH₃ in Ration KTSPSR. One possible reason to the increase of NH₃ concentrations is the use of vitamins which may positively impact ruminal protozoa and aiding in the enhancement of ammonia nitrogen levels in the rumen fluid (Naziroglu et al 2002; Silva et al 2022). However, the precise mechanism through which vitamins or minerals might augment the concentration of ruminal protozoa is not covered in our study.

However, the diverse feeding rations and the addition of feed supplement did not affect CH₄ gas production (p>0.05) but there was a tendency for decreased CH₄ gas production in KUDS. The decreased in CH₄ gas production can be attributed to the pH reduction. The decrease of pH has suppressed the activity of methanogenic archaea, leading to a subsequent reduction in methane production in the rumen (Schroeder et al 2014). The feed supplement in the diverse feeding rations successfully increased total VFA. Feed supplement has fulfilled the dairy cow's mineral requirements, including iron, copper, manganese, selenium, cobalt, and iodine, as well as their vitamin needs, such as A, D3, E, Hy D, B12, biotin, and niacin. Similarly, Astawa et al (2011) achieved a comparable increase in VFA production when using vitamin-mineral supplement in commercial feeds. In addition, the feed with the lowest energy and protein content resulted in the lowest total VFA concentration (KUDS).The discrepancies in the existing literature concerning the impact of vitamins and minerals supplementation may be attributed to differences in environmental conditions, animal categories, and other influencing factors (Schafers et al 2018). The VFA values in the study remained within the range necessary to support microbial growth in the rumen, as outlined by McDonald et al (2022), which is typically between 70-150 mMol. The results demonstrated an interaction between the diverse feeding rations and feed supplement.

Table 8 presents the proportion of partial Volatile Fatty Acid (VFA) concentration values. The diverse feeding rations and feed supplement did not affect (p>0.05) the partial VFA concentration (C2, C3, C4 nC4, iC5, and nC5).

The findings of the study has revealed the significance of using feed supplement to increase feed efficiency and to potentially mitigate greenhouse gas (GHG) emissions in dairy farms. Nevertheless, the use strategies to use feed supplement is hindered by the substantial cost incurred by dairy farmers and a knowledge gap regarding the importance of feed supplement. To address this challenge, financial initiatives aimed at bolstering feed efficiency through the implementation of feed supplements can be implemented. Additionally, it is crucial to educate farmers about the importance of utilizing feed supplements.

Conclusion

In this study, we evaluate the effects of feed supplement within diverse dairy cattle feeding rations on in vitro fermentability, ration digestibility and methane gas production. Based on our study, we found that different feeding rations and feed supplement impacted rumen pH, the concentration of N-NH₃ in the rumen, the total VFA concentration, and dry matter and organic matter digestibility but did not affect the CH₄ gas production. In addition, we also identified farm management systems and highlighted the variability in feeding management systems across the different farms. At the farm level, the KUDS diet was better than others due to the high protein feed offered to the cows, accompanied by higher dry matter and organic matter digestibility but yielded a high N content in faeces. Given the context of Foot and Mouth Disease (FMD) outbreaks, maintaining milk yield emerges as a crucial concern. During the FMD outbreaks maintaining the milk yield is necessary and providing good quality of feed, which encompasses the use of feed supplement, holds promise as a viable strategy for the dairy farms facing such challenges.

Conflicts of Interest

The authors declare that they have no conflict of interest.

Authors’ contributions

ZWA conceptualized, implemented the experiment, curated the research data, and wrote the original draft; IGP conceptualized and designed the experimental, reviewed, and edited the manuscript draft; SS conceptualized and designed the experimental and edited the manuscript draft; AJ reviewed, and edited the manuscript draft; AI research material preparation, curated the research data and edited the manuscript draft; AS research material preparation, curated the research data and edited the manuscript draft; and WWW designed and conceptualized the research and edited the manuscript draft.

Acknowledgments

The authors would like to thank the PT. Nutricell Pacific for the research funding.

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