The current study was focused on the disparity of two methodologies to deepen our perception of the digestible value of 14 feedstuff tested for use in the practical diet of sub-yearling beluga and the fecal collection method's approach to assessing their digestible value. Before the dietary fine-tune formulation, the ascertainment of ADCs of feedstuffs is an imperative indicator to detect nutrient bioavailability (Allan et al., 2000; Jannathulla et al., 2018; Mirzakhani et al., 2020).
Fish meal apparent digestibility
In the present study, the ADCCP values of fish meal class in settlement and stripping methods ranged from 89.4-94.9% to 83.6-87.7%, respectively, which denotes that fishmeal feedstuffs were digested efficiently by beluga. In accordance with our result, Safari et al. (2016) observed that the digestibility of Anchovy protein in settlement and stripping methods in subadult beluga was 93.2% and 86.1%, respectively. In the other study, the authors addressed that the highest (92.9%) digestibility of fishmeal (Clupeonella sp.) in stripping method was observed in the yearling Siberian sturgeon (Mirzakhani et al., 2020). Our findings revealed that the fishmeal class was significantly more digestible than animal protein and botanic origin. The priority of ADCCP in AM and FFAM compared to other fishmeal class could be attributed to the alterations in protein solubility and dispersibility attributes (Jannathulla et al., 2018; Mirzakhani et al., 2020). Studies including various fish species have declared the superiority of fishmeal due to, in part, its well-balanced essential amino acid content (not measured in our study) and lack of anti-nutrient factors (Allan et al., 2000; Liu et al., 2009; Mohanta, Mohanty, Jena, & Sahu, 2006; Safari et al., 2016; Tibbetts, Lall, & Milley, 2004). In addition to appraising the digestibility of fishmeal, the freshness and type of the raw ingredients and processing during manufacture need to be taken into account.
There is a consensus that the high values of indigestible substances such as ash in feedstuffs could have reduced gastric retention time and increased evacuation time through the gastrointestinal tract (Colombo et al., 2020; Guerrero-Zárate et al., 2019; Mirzakhani et al., 2020; NRC, 2011). This event and the low quality of the raw material or processing conditions could be the underlying causes of the inferior ADCCP of the TBM than the other fish meals in our study.
The digestibility of dry matter gives overall evidence of feedstuff digestibility, indicating the digestible fraction of inorganic and organic material, which are deeply corroborated to their ash and insoluble carbohydrates such as fiber (Basto et al., 2020; Brunson et al., 1997; NRC 2011). In line with this, our work demonstrated an increase in ash content accompanied by attenuation of ADCDM in TBM.
Our outcomes revealed that ADCCP of fishmeal class in settlement method was numerically 5.9% higher than stripping method. A plethora of literature announced that the stripping event has the lowest ADCs compared to the settlement or column method (Blyth, Tabrett, Bourne, & Glencross, 2015; Glencross, Booth, & Allan, 2007; Heinitz et al., 2016; Safari et al., 2016; Vandenberg & De La Noüe, 2001). Vandenberg and Noüe (2001) postulated ADCCL values fluctuate to a lesser degree when two collection methods are used. They attributed the hydrophobic nature of the solubility of feedstuffs in diet, leading to inferior nutrient leaching.
Animal protein apparent digestibility
In the present study, relatively low digestibility coefficients of macronutrients were recorded for animal by-products components. Several studies have declared that PBM is a promising rendered animal protein feedstuff for sturgeon diets. Liu et al. (2009) postulated that ADCCP and ADCDM of PBM in the basal diet using a ratio of 70:30 in the juvenile Siberian sturgeon, Acipenser baerii, in the fecal collection method were 86.1% and 59%, respectively. While these values, in our study, were 81.54% and 77%, respectively. In our previous study, outcomes revealed that juvenile beluga did not show decreasing trend in ADC of overall macronutrients in the fecal stripping method up to 60% substitution of FM by PBM (Sayed Hassani et al., 2021). In the other study, Mirzakhani et al. (2020) stated that ADCCP in PBM in the stripping method was 83% that was 5% more than our study in the same procedure. This variety could be due to differences of crude protein and ash in the PBM in two experiments. The heterogeneity of nutritional value of PBM reported by numerous authors may be due to different origins and processing methods (Shapawi, Ng, & Mustafa, 2007), presence of a high level of feather meal that leads to poorly digestible keratin protein (Tibbetts et al., 2011), inferior indispensable amino acid and presence of disulfide bonding (Moran et al., 1966) and the residence of indigestible amino acid in adverse processing method, vis. lysinoalanine and lanthionine (Mirzakhani et al., 2020; Tibbetts et al., 2011). As with earlier studies in sturgeons (Liu et al., 2009; Safari et al., 2016), the lowest digestibility of protein, lipid, and dry matter in animal protein was observed in the BM. Meanwhile, Allan et al., (2000) acknowledged that dry matter, protein, and energy digestibility in the Australian silver perch, Bidyanus bidyanus, included in the reference diet (with a 69: 29 ratio) was 109%, 90.2%, and 99.9%, respectively. These results could explain the more leakage of faces collection, differences of fish species, degree of protein hydrolysis and BM processing (Alarcón, Moyano, & Díaz, 2002; Lee, 2002).
It is also well established that the MBM have inferior ADCs for lysine than the other marine or animal by-products meals. This could indicate heat processing leads to lysine degradation (Allan et al., 2000; Zhou et al., 2004). Also, the presence of fibrous components of bone in this feedstuff, together with unsuitable processing, could have lowered the protein digestibility by disulphide bridges, cross-linkages, denaturation of protein, especially lysine, and lowering gut evacuation materials and physically shielding protein from digestive enzymes (Allan et al., 2000; Jobling, Gomes, & Dias, 2007). In agreement with our study, Liu et al. (2009) declared that the digestibility of crude protein (86.1%) in MBM, and especially for lysine and methionine (82.1% and 84.9%, respectively) by fecal collection method in the juvenile Siberian sturgeon lower than the other tested feedstuffs.
Plant origin apparent digestibility
Our finding revealed that processing plant feedstuffs (WG, CG and PSM) due to removing the crude fiber and anti-nutrient factors had a higher overall digestibility of macronutrients in sub-yearling beluga. In corroborated with the other studies, ADCs of dry matter and gross energy were related to the carbohydrate content (Allan et al., 2000; Safari, Shahsavani, Paolucci, & Mehraban, 2014; Tibbetts et al., 2004). Processing plant proteins can notably quench the antigenicity of these feedstuffs in the aquaculture feed industry (Drew, Borgeson, & Thiessen, 2007; Hosseini Shekarabi, Shamsaie Mehrgan, Banavreh, & Foroudi, 2021), which was in accordance with the results obtained in two types of SM in our study. In stripping and settlement methods, the ADCs of macronutrients of PSM was roughly 4-5% higher than SM. Liu et al. (2009) declared that juvenile Siberian sturgeon showed effective protein digestibility on SM in the collection method (91.8%), while a drastic reduction was observed in ADCCP of full-fat SM in subadult beluga (51.1%) by fecal collection method (Safari et al., 2016). This heterogeneity could be attributed to the different species, fish age, various production processing, levels of anti-nutrients and origin varieties. The alleviating of digestibility of crude lipid in SM may be due to the binding of bile acid, reducing lipid metabolism, which is in line with Dalsgaard et al. (2012). The type of diet (pellet and extrude) is very important in the digestibility of macronutrients that need to be taken into account. One of the most pronounced reduction ADCs was related to the CAM. The explanation would be that, in part, to present non-starch polysaccharides (NSPs) as well as anti-nutrient factors such as glucosinolates and phytic acid that could be attenuated the ADCCP in the test diet (Gatlin et al., 2007; Kaiser, Harloff, Tressel, Kock, & Schulz, 2021; Danwitz & Schulz, 2020). A plethora of studies on fish indicates that the ADCGE value in CAM is inferior to those found for fish meals (Lim et al., 1997; Safari et al., 2016).
Lee (2002) also well established that ADCs of dry matter and gross energy of plant feedstuffs are associated with the content of carbohydrates, so that an increase of NSPs in the diet was negatively related to the digestibility of the two macronutrients mentioned above, which was in accordance with our study. In other words, it can be claimed that carnivorous fish tend to utilize the dry matter and energy in animal proteins more beneficial than that in plant origin feedstuffs.
Settlement feces of collection vis-a-vis stripping method in apparent digestibility
It is acknowledged that the type of collection of feces appears in various digestibility evaluations (Glencross et al., 2007; Rawles et al., 2010). In the current study, the ADCs of feedstuffs in the stripping method were 1-15% lower than the settlement method, which relatively concurs with the other studies (Glencross et al., 2007; Safari et al., 2016). It should be noted that the domain between the two methods in different studies could be related to the production of diet (extrude vs. pellet) and the composition of feedstuffs. Intuitively, however, despite the loose stability of feces in the sturgeon species, this can be one of the reasons for the greater difference between the two collection methods in this species. As shown in the results, the most significant differences in the ADCs of dry matter, crude lipid, and gross energy were recognized in plant-origin feedstuffs. A plethora of studies lubricates that the presence of a high value of carbohydrates in the feces reduces the integrity of feces in the passive collection method, upon which fecal matter dissolved in water media. Explaining this phenomenon could occurrence of nutrients leaching associated with exposure time in the settlement technique, leading to overestimation in apparent digestibility (Blyth et al., 2015; Glencross et al., 2005; Glencross et al., 2007; Rawles et al., 2010). Contrarily to this prospective, feces evacuation by hand in the stripping method over the hindgut, leading to the expulsion of the intestinal epithelium, digesta (the diet with incomplete digestion), mucus, bile, urine, enzymes, and blood, resulting in skewed results and hence underestimation of digestibility (Lee, 2002; Percival, Lee, & Carter, 2001; Storebakken et al., 1998). Generally speaking, stripping provided more conservative digestibility of diet/feedstuff than those achieved applying for the settlement procedure.