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Text|Broken Boat Han
Editor|Broken Boat Han
preface
As aquaculture expands, aquaculturists are increasingly aware of the need for appropriate strategies to reduce farming costs and improve feed efficiencyToday, the application of feed supplements is particularly popular among fish farmers because of their positive impact on fish growth and health.
Acidulants such as lactic acid (LA), formic acid, acetic acid, propionic acid, and citric acid are examples of feed supplements used in fish diets, where acidifiers play a role in the growth and development of intestinal villi and increase the absorption surface.
In addition to this, various studies have shown that these dietary supplements are used as nutrients by beneficial gut bacteria, thereby increasing the number of these bacteria and ultimately lowering the intestinal pH.
In addition to promoting the elimination of pathogens, the increase in intestinal acidity leads to the dominance of beneficial microorganisms, which secrete digestive enzymes and degrade anti-nutritional factors for better digestion and absorption, and acidulants effectively increase the secretion of intestinal mucin.
In addition to keeping digestive cells healthy from feed scratches, it can also increase pathogenic bacteria, washing organic acids from the fish digestive system, effectively reducing oxidative stress and increasing antioxidant capacity, thereby improving fish health and welfare.
There are also acidulants that stimulate the immune system of fish and increase the expression of some immune genes, such as lysozyme (lys), interleukin-1β (IL1b), tumor necrosis factor-α (TNFA) and defensin (def) in fish disease resistance.
LA is one of the well-known acidulants in the aquaculture industry and is used as a feed supplement, and studies have shown that diets containing 10-20 g/kg LA can improve the growth performance and digestibility of nutrients in beluga whales, as well as improve immunological and antioxidant markers in rainbow trout.
The addition of sodium lactate to diets improved growth performance in narrow-clawed crabs and Arctic sockeye salmon, but LA and its salts had no effect on weight gain and feed efficiency in rainbow trout and Atlantic salmon, suggesting that further research is needed to illustrate the effects of LA on fish.
Materials and methods
Fish feed is made using locally available feed and according to the nutritional needs of carp, the feed is weighed and mixed, then 400 mL/kg of water is added to the mixture to make the dough, and LA is added at the expense of 2.5 g, 5 g and 10 g/of water.
Diets not supplemented with LA are considered control diets, meat grinders are used to make feed pellets, dried on fan blowers, crude protein (Kjeldahl method), crude fat (ether extraction), crude ash (burned in a muffle furnace), and crude fiber (acid/alkali digestion) in the diet according to standard methods.
The intestinal samples were homogenized with three volumes of phosphate buffer in a mortar and in the presence of liquid nitrogen, and after homogenization for 2 min, the homogenate was poured into three separate 2 mL tubes
Total soluble protein was measured using the Bradford method, and as previously reported, trypsin activity was determined at 410 nm using DL-arginine-p-nitroaniline as a substrate, and N-benzoyl-L-tyrosine ethyl ester was used as a substrate for the determination of chymotrypsin activity at 256 nm.
Amylase activity is determined according to the method described earlier, using soluble starch as a substrate, the increase in the reducing power of the buffered starch solution is measured at 540 nm with 3-5 dinitrosalicylic acid (DNS), lipase activity is based on hydrolysis of p-nitrophenyl myristate at 37 °C and wavelength 580 nm.
Alkaline phosphatase (ALP) activity is determined for p-nitrophenol and phosphate based on p-nitrophenyl phosphate, hydrolyzed in the presence of magnesium ions, as described previously.
Organic acids are gaining popularity in aquaculture as feed additives because of their advantages such as improved nutrient digestibility, growth performance, immune stimulation and gut health.
The effectiveness of diets supplemented with organic acids in promoting fish growth performance is not always consistent, and studies have shown that the use of LA or its salts can lead to different results, for example, the addition of 20 g/kg LA to fishmeal and plant-based diets improves beluga whale growth performance.
Supplementation with 16.7g of sodium lactate did not promote the growth performance of giant grouper and willow leaf algae fed on a plant-based diet, and when fed a diet containing 5-20g/kgLA, rainbow trout had no growth-promoting effect, in crustaceans.
Supplementation with 5-50 g encapsulated sodium lactate significantly improves the growth performance of narrow-clawed crayfish, and these inconsistencies may be due to species-specific characteristics, to support this, the findings comparing Arctic salmon and Atlantic salmon under similar experimental conditions were found.
Dietary supplementation with sodium lactate is beneficial for Arctic salmon but not for Atlantic salmon, a difference that experts attribute to Arctic salmon's longer gastric emptying time, which reduces harmful bacterial populations in the gut.
One of the benefits of organic acid supplements is their ability to improve nutrient digestibility, which can improve the growth performance of fish.
Similarly, LA supplementation has been shown to increase pancreatic enzyme activity in fish gut, as shown in previous studies of other organic acid-treated fish.
Studies have shown that organic acids increase the activity of pancreatic enzymes in fish intestines, but it is not clear whether this increase is due to the secretion of pancreatic enzymes into the lumen, or whether they come from microorganisms in the lumen.
Experts say that an increase in the source of plant protein in the fish diet will reduce the activity of digestive enzymes in the intestines, but not the activity of the liver and pancreas, and organic acids may not have a significant effect on pancreatic digestive enzymes.
For example, in shrimp, pancreatic enzymes are sensitive to the antinutrients in plant proteins, in which case intestinal digestive enzymes are significantly reduced.
For example, phytates are well-known antinutrients in plants that inhibit lipase, amylase, and protease activity, beneficial bacteria can secrete digestive enzymes in fish intestines, and they can degrade antinutrients and inhibit their negative effects on digestive enzymes.
For example, many lactic acid bacteria produce phytase, which breaks down dietary phytates and may improve the activity of digestive enzymes.
It is hypothesized that dietary LA increases the intestinal activity of amylase, lipase, trypsin and chymotrypsin by altering the intestinal microbiota.
Experts have also found that LA supplementation increases intestinal ALP activity, a brush-like boundary enzyme responsible for nutrient absorption.
Elevated ALP may be due to increased enzyme production and/or intestinal folding, resulting in an elevated intestinal surface weight ratio, and some articles have reported results that support the claim that organic acids increase intestinal folding in fish.
Dangerous events
Oxidative stress is one of the most life-threatening events in fish, because the concentration of unsaturated fatty acids in the fish is high, and the increase in pro-oxidants in the fish body leads to the activation of the antioxidant system, characterized by increased activity of antioxidant enzymes to neutralize harmful compounds.
If the antioxidant capacity does not completely remove harmful substances, they attack biomolecules, especially fatty acids, which are the products of lipid peroxidation.
Low MDA concentration is an indicator of low oxidation state, and GSH-dependent antioxidant factors play an important role in protecting fish cells from oxidation.
GPx uses GSH as a cofactor to neutralize hydrogen peroxide and other hydroperoxides, and as a result, glutathione is oxidized and loses its biological function.
GR is responsible for reducing oxidized glutathione, and dietary LA supplementation increases GSH levels and decreases GPx and GR activity while lowering MDA levels.
LA improves the antioxidant capacity of the gut, resulting in decreased glutathione oxidation, which reduces the activity of glutathione-related enzymes, and dietary organic acids/salts have been reported to increase the antioxidant capacity of fish.
While data on this topic are limited, dietary LA supplementation has been shown to significantly increase antioxidant enzyme activity and reduce MDA concentrations in fish, and the antioxidant modulation effects of dietary LA supplementation may be associated with improved fish gut microbiota.
Beneficial bacteria can enhance the antioxidant capacity of fish, specifically, some lactobacilli are able to produce GSH and riboflavin.
The efficacy of organic acids
This is required for the GSH redox cycle, so it is speculated that the increase in lactobacillus populations in fish intestines increases the antioxidant capacity of fish intestines and reduces lipid peroxidation.
Organic acids can alter the gut microbiota by lowering gut pH and acting as nutrients for certain bacteria, and after utilizing specific organic acids/salts, host gut microbes release other metabolites that act as nutrients for other species, potentially altering the entire microbiota of fish gut.
Experts believe that dietary butyrate, acetate or LA increases the number of lactobacilli in fish intestines, which is consistent with current results, dietary LA supplements subordinate populations of harmful bacteria.
This is consistent with previous studies showing that organic acids reduced Streptococcus suis, Pseudomonas and Vibrio in fish intestines.
So dietary LA supplementation may be an effective procedure for balancing the gut microbial population of carp, which may be the main reason why LA is beneficial for this species.
The gut plays a vital role in the immune function of fish, because it is a barrier against harmful pathogens, and the interaction between intestinal flora and antigens and host fish occurs on the surface of the mucosa of the gastrointestinal tract, and fish rely on defensins to achieve various immune-related functions.
For example, antibacterial, antiviral and anti-inflammatory effects, dietary feed additives upregulate intestinal expression, known as the improvement of fish intestinal immunity.
There are few data on the effect of organic acids on fish gut expression, and dietary LA supplementation has no significant effect on def expression in rainbow trout gut.
Dietary LA administration is an effective and practical way to increase intestinal expression in carp and may help enhance intestinal immunity and health, and shock protein 70 is generally thought to protect living cells from stressful conditions by maintaining proper protein folding and function.
The expression of hsp can vary under different conditions, for example, treatment stress does not affect the HSP expression of rainbow trout, but reduces the gene expression of yellow bass, and similarly dietary LA and malic acid supplements can upregulate the expression of intestinal HSP in the intestine of rainbow trout.
These upregulation of intestinal HSP expression is consistent with the innervation of beneficial bacteria or enhances the antioxidant capacity of fish gut, and it is assumed that the increase in intestinal HSP expression in LA-treated fish is a positive response that enhances the cell's ability to process potential stressors, suggesting that upregulation of HSP expression is a sign of enhanced gut health.
Fish has a variety of glycoproteins called mucin, which are present in the mucosal barrier, of which mucin 2 and mucin 5 are important macromolecules that produce the gel layer, and studies in humans and broilers have shown that short-chain fatty acids may induce signals to increase mucin.
Dao mucin is essential for the immune response and changes during infection or exposure to toxic substances, and it has been found that fish enteric mucin depletes within the first few days after a bacterial/parasitic infection which is a defense mechanism for clearing pathogens, but recovers afterwards.
Immune cells produce cytokines such as IL1B and TNFA, which initiate the immune system's defense mechanisms by enhancing phagocytosis, respiratory burst activity, and nitric oxide production, lysozyme is an essential molecule for innate immunity and has powerful catalytic power.
Can decompose bacterial cell wall, promote cell lysis in low osmotic environment, lysozyme has antibacterial, antiviral and anti-inflammatory properties, can activate nuclear leukocytes and macrophages, promote phagocytosis, lysine, IL1B and TNFA expression levels, is an important indicator to monitor fish immunity and evaluate the effect of diet and vaccine.
Similar to the current results, dietary malic acid and butyric acid increase the expression of lysates, il1b and TNFA in fish intestines, and it has been found that the inducing of lysate, IL1b and TNFA expression in fish intestines after administration of organic acids is accompanied by higher disease resistance.
Interestingly, this increase in disease resistance, accompanied by the domination of beneficial bacteria and the subordination of harmful bacteria in the intestines of crucian carp, may partly explain the current results.
conclusion
Dietary lactic acid supplementation improves fish growth performance and feed efficiency by enhancing intestinal enzyme activity, regulating gene expression associated with immune responses, increasing beneficial bacterial populations, and providing antioxidant benefits.
It appears that these benefits are mediated by the domination of beneficial bacterial communities in the gut, and although most test parameters indicate that 5g/kgLA is suitable for carp dietary supplements, further research is needed to address the reasons behind differences in response of certain parameters to dietary LA levels.
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