1. Introduction
China is one of the largest producers in agricultural production, and the annual crop straw production was approximately 810 million tons in 2013 [
1], of which 190 million tons was rice straw [
2]. Rice straw is the major source of biomass yield, and rice grain is distributed throughout the globe because of its importance as a kind of food crop for more than half of the world’s population [
3]. The increasing consumption of grain leads to a large amount of straw residue, including rice straw with low utilization. The low utilization rate of rice straw is mainly because of its low nutritive value, which is commonly considered as a medium-low quality feed in dairy cattle rearing [
4,
5]. Therefore, concerns in improving the nutritional quality and utilization rate of rice straw by ruminants are stayed increasing.
So far, various preprocessing methods have been explored in practice, such as physical, chemical and biological treatment to enhance the nutritional quality of straw [
6]; ammoniation is the most key chemical technique among them [
7]. Ammoniation has been developed to break lingo-cellulosic bonds in crop residues, thereby enhance the nutritional quality of rice straw [
8]. Urea is a commonly used chemical product in ammoniation treatment [
9]. However, it has been reported that urea treatment only retains 30% to 35% of the NH
3 released during the treatment [
10]. Feeding cows the urea treated rice straw, the retained nitrogen would rapidly liberate in the rumen and cause nutrient losses [
11]. The release rate of urea in the rumen is too fast, and the available nitrogen is much higher than the requirement of energy and nitrogen balance. At present, many researchers are focusing on reducing the release rate of nitrogen in the rumen to achieve the effect of energy and nitrogen balance [
12]. To tackle this problem, some researchers utilized H
2SO
4 or HCl to fix NH
3 [
8]; however, fixing the NH
3 in straw with acids was costly and dangerous. The addition of urea can increase the total nitrogen content in the feed, but with the release of ammonia, the pH value increases, resulting in the fermentative quality of the silage declined [
13].
Corn steep liquor may solve the problem of escaping NH
3 of urea treated rice straw. It is a kind of by-products of processed corn starch, rich in carbohydrates and can significantly increase the crude protein (CP) content of rice straw [
14]. Corn steep liquor has an ability to fix NH
3 because of its acidic characteristic [
15], which increased the CP content in the rice straw. Nisa et al. studied the effect of supplementation of 0%, 3%, 6% and 9% dry matter (DM basis) of corn steep liquor to wheat straw treated with 5 kg urea (50% moisture level) [
16]. A higher retained nitrogen content and improved digestibility of DM and neutral detergent fiber (NDF) were found in the diet containing corn steep liquor of Buffalo bulls. In addition, as the concentration of corn steep liquor increased, the rumen volatile fatty acid (VFA) and acetic acid (AA) concentrations increased, while the highest VFA concentrations were found with the addition of 9% corn steep liquor [
15]. As a feed flavor additive, molasses could enhance the palatability [
17] and increase the feed intake of ruminants. Furthermore, it has been reported that molasses improve the quality of silage fermentation [
18]. However, most of the previous studies have not considered the combined effect of urea, corn steep liquor and molasses on the nutritive value and digestibility of rice straw.
Therefore, the aim of the study was to explore the effects of different ammoniation treatment methods on the nutritional quality parameters (chemical composition, in vitro digestibility, in vitro gas production and in vitro rumen fermentation performance) of rice straw. In addition, the current study evaluated the process to provide a scientific foundation for the utilization of rice straw as feed.
4. Discussion
Straw is a common source of feed for ruminants, but its utilization efficiency is low due to the low CP content with only 3~5% of DM and high NDF and ADF [
25]. In the present study, no difference was found in the DM of rice straw for all groups, which showed that the ammonia treatment has little effect on the DM content. Furthermore, the nitrogen content of feed affects the efficiency of rumen microbial protein synthesis, and low nitrogen utilization ability will cause economic losses and environmental pollution. Our current findings revealed that 9C5U and 9C2.5U improved the CP levels of rice straw from 5.8% to approximately 11.0% after 60 days of anaerobic storage. Similar to our results, a study has shown that the nitrogen content increased in rice straw by the acidification of corn steep liquor [
26]. Furthermore, with the addition of molasses, the nitrogen retention rate in rice straw was reduced, which might be due to the molasses and urea reaction [
18]. The addition of corn steep liquor could increase the nitrogen retention rate in the rice straw which is why the CP content in 9C5U and 9C2.5U is higher than 5U treatment. The main reason for this might be due to the losses of ammonia in 5U treatment. Generally, a higher CP level is beneficial for higher DM digestibility [
27]. This is noteworthy that in the present study, the corn steep liquor treatment improved the CP content of rice straw. Corn steep liquor promotes the proliferation of microorganisms by providing excellent carbon and nitrogen substrates metabolism, thereby increased the synthesis of functional enzymes and bacterial proteins and improved the CP content of rice straw [
28]. In general, another reason for the low utilization rate of rice straw is the high content of NDF and ADF, which decreases the digestibility in ruminants. The 9C5U treatment decreased NDF and ADF of rice straw from 65.41% to 60.02% and 54.64% to 47.84% after 60 days of anaerobic storage. This showed that ammoniation treatment caused the breakdown of the complex structure of rice straw lignocellulose and dissolved a part of cellulose and hemicellulose [
29]. Consistently, the ammoniation treatment ensures the sufficient nitrogen supply of microbes, which resulted in the higher degrading activity of the microbes and the resulting effects.
DM and NDF digestibility are the key characteristics that stand for the intake and production potential of forages in ruminant feeding. It was noticed in our findings that all the treatments improved IVDMD and IVNDFD of the rice straw after 60 days of anaerobic storage. Ammoniation weakens the hydrogen bonding inside the rice straw; result in the expansion of fiber molecules and the ester bond or ether bond. Bacteria are the main settlers in the rumen and therefore make the most significant contribution to the fermentation, degradation and digestion of feed. It was documented in previous findings that the barley straw fibers treated with urea cultured in the rumen of animals have the most bacteria attached and the change of rumen environment significantly increased the degradability of rumen bacteria [
30,
31]. The 9C5U treatment was most effective which may be due to the addition of the highest quantity of urea and corn steep liquor. The level of IVDMD and IVNDFD in 9C2.5U and 9C2.5U3M treated group was lower than the 9C5U treated group, the less quantity of the urea addition in 9C2.5U and 9C2.5U3M group might be the possible reason for these differences. However, the IVDMD and IVNDFD of the 5U group were lower than that of the 9C5U group because of no corn steep liquor addition, which makes the rumen microorganism obtain less nutrients and shorts the rumen fermentation [
32]. There was no statistical difference among 5U, 9C2.5U and 9C2.5U3M. Moreover, in the corn steep liquor-added group, the IVDMD and IVNDFD were higher than the control group. We expected that differences might be due to the rapid and simultaneous release of corn steep liquor and urea to a certain extent, which increased the growth efficiency and the number of microorganisms [
14,
33]. On the other hand, the DM degradation rate can reflect the digestibility of the feed to a certain extent. It is related to the nutritional composition of the feed. Generally, a higher degree of synchronization of energy and nitrogen release increases the digestibility of feed nutrients in part by affecting the bacterial community, metabolism and enzyme activity of ammonia assimilation in vitro fermentation [
34].
During in vitro fermentation, the digestibility of feed organic matter is significantly correlated with gas production. The higher the digestibility of organic matter, the fermentation activity of microorganisms in the rumen increases, the gas production rate is accelerated, and the gas production is further increased [
35]. In the present study, the GP
48 and AGPR of each ammoniation treated groups were higher than that of the control group, indicating that the ammoniation can increase the soluble carbohydrate of rice straw and increase the gas production rate and the gas production. It was noticed that chemical treatment changed the rice straw cell wall structure and the solubility of cellulose and hemicellulose is improved [
36], which can be explained by significant differences in NDF and ADF content. The results of in vitro studies further indicate that the higher the urea content, the greater gas production in vitro, which is consistent with the results of the Senthilkumar’s findings [
37]. The GP
48 in the 9C5U and 5U groups were the highest and was consistent with their low NDF and ADF content. In the 9C2.5U and 9C2.5U3M groups, the urea content was reduced and the NDF and ADF content was higher, which may lead to lower gas production in vitro, But they are better than the control group.
The pH value of the treatment groups in this study was not significantly different within the normal range from 6.75 to 6.82, which was consistent with the report by Bath et al. [
38]. This shows that the effect of different compound ammoniation methods on the rumen pH was not obvious in this study. The AA, PA and BA were increased in the compound ammoniation treatment groups compared with the control group, which may be due to the enhancement in the digestibility of rice straw [
39], making it more degradable, and increased VFA production. Among them, the contents of AA, PA and TVFA in the 9C5U and 5U treatment groups were the highest, which was consistent with their highest gas production in vitro. The 9C5U, 5U, 9C2.5U and 9C2.5U3M treatments had a higher PA level than the control. A previous report had documented that the PA production was positively correlated with nitrogen deposition, and the higher CP content in these treatments than control also explains this phenomenon [
40]. Besides CP, the microbial supply and activity were also enhanced by ammoniation treatment which is also one of the leading factors responsible for the higher level of PA.
Ammoniation treatment degraded the side chains of esters and glycosides leading to structural modification of lignin, cellulose swelling, cellulose decrystallization, and hemicellulose solvation and increased the degrading activity of the microbes [
41], and this diversification might affect the NDF and ADF of rice straw. In the current study, the NDF and ADF content of rice straw and IVDMD and IVNDFD also showed a negative correlation. In addition, the CP content in the straw was positively correlated with digestibility, gas production and rumen VFA content [
42], which are in line with our current findings.
It was previously reported that rice straw treated with urea (50 g of urea per kg DM straw) was used to replace elephant grass to feed lactating cows. It was found that the use of urea-treated rice straw increased the replacement amount of elephant grass until 75% of the roughage. Furthermore, rice straw had increased the milk fat content, and has no effect on milk yield and other milk composition parameters. Feeding rice straw treated with urea could partially replace the elephant grass in the lactating cow’s diet during the dry season, which reduced the cost of roughage [
43]. In addition, feeding rice straw treated with urea enhanced the rumen ecology, rumen fermentation efficiency and nutrient digestibility of ruminants [
44]. Studies have shown that rice straw treated with 20 g/kg urea + 20 g/kg calcium hydroxide can increase nitrogen retention and microbial protein synthesis [
45].