Part IV: Nutrients

Proteins
Proteins are organic compounds made of amino acids linked to one another through peptide bonds. Proteins are also first choice of energy source for Koi because the net energy they gain from protein is far higher than from fats and sugars. Amino acids contain “amino groups’ that are formed from Nitrogen and Hydrogen. The nitrogen content of proteins found in animals is about 16%. This is important in calculating the ammonia produced by Koi when given a feed containing certain percent of protein. After ingestion, the proteins are broken down into amino acids and released into blood. Tissue cells absorb these amino acids from blood and use them to build/repair new tissues.

If there is excess protein in the feed, Fish can not store them. Instead, they use them as energy source, over sugars and fats, for movement and staying suspended in water. So, if you feed high amounts of protein, fish does not grow exponentially. It will  use the excess protein to produce energy to explore the pond. As fish grow, they need less protein because there is no more tissue building.

Wilson and Halver conducted an experiment with Common carp in 1986 and found that protein requirement decreases over age. Feed for fry and fingerlings must contain 50% protein. In salmon, this requirement decreased to 40% in 6-8 weeks and to 35% for yearlings raised at standard environmental temperature.

But, appetite and efficiency of digestion depend on water temperature. They increase proportionally with water temperatures until water temperature reaches 80s. Growth rate, however, increases  at a faster rate because fish eat more as temperature increases and what they eat, gets converted into tissue growth more rapidly at higher temperatures.

Gross protein requirement for normal growth of Carp is 380g/kg. The following 10 amino acids are Essential for normal growth of Koi but Koi can not synthesize them in their body. So, we must provide them in the feed.
arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine

Feed containing fishmeal, bone-meal, meat, yeast, soybean can be improved with the addition of cystine (10g/kg) and tryptophan (5g/kg).

Fats
Fats are energy deposits for Fish. Even though Koi can extract more energy from proteins, they do not store protein as energy depot, So, when there is need for extensive energy like during starvation, or spawning, Koi breakdown the stored fat rapidly, to produce energy. Because of this unique ability, they are able to stay without food throughout winter even though they are warm water fish. Fish contain lots of unsaturated fats. Unsaturated fats contain double bonds at certain positions in their chains. The w number identifies the position of the first double bond in the chain from the methyl end. If there are many unsaturated bonds, then the fatty acid is called Poly Unsaturated Fatty Acid or PUFA. Unsaturated fatty acids are “hungry” meaning they want to saturate themselves to get rid of the double bond to become more stable single bond. So, as soon as they are exposed to air, they oxidize themselves. In the process they go rancid!

Unsaturated fats have lower melting points (meaning they remain in the liquid form at lower temperatures whereas saturated fats like butter turn into solids. Homothermic animals like humans have the ability to heat themselves. So, saturated fats remain in the liquid form in the blood. But temperature of fish is dependent on the water temperature. So, if they have unsaturated fats, they can remain in the liquid form in the blood at lower temperatures.

Experiments have shown that Carp need both w 3 and w 6 fatty acids. The best weight gain and growth was seen when the fish were fed food with 1% each of 18:2 w 6 fatty acid and 18:3w 3 fatty acid. Also,  20:5w 3 fatty acid and 22:6w 3 fatty acid at 0.5% of the diet was more effective than 1% of 18:3w3 fatty a cid. The need for unsaturated fats in the diet is biggest problem in manufacturing and storage of Koi food because food can go rancid quickly. Rancidity can result in some products that can interact with proteins and vitamins and make them ineffective. Some of the oxidized products are toxic to Koi. So, it is extremely important to buy fresh food and store them in air-tight, light-proof storage jars in a cool, dry place. Scientists have observed dark coloration, anemia, lethargy,  brown-yellow pigmented liver, abnormal kidneys, and small gill clubbing  in chinook salmon, fed with rancid food. if food contains high amount of unsaturated fats, adequate amounts of Vitamin E (an anti-oxidant) should be added to the food.

Carbohydrates
Sugars, starches, celluloses and gums are called carbohydrates. Carbohydrates are the main energy source for homothermic animals (like proteins for fish). About 75% of plant biomass is carbohydrates but animals have only stored glycogen and some parts of tissues that are carbohydrates. Carbohydrates are polymers of sugars. So, no matter what form of carbohydrates are consumed, they need to be broken down to the simplest sugar form i.e. glucose in order to be used. Most animals, including fish, are not able to digest plant starch (cellulose). But nearly all herbivores including Carp have microorganisms living in their gut that are capable of digesting cellulose. Legume seeds and soybean contain significant amounts of  carbohydrates as medium-size polymers of a sugar called galactose rather than glucose. Fish do not have enzymes to breakdown these oligosaccharides. If soybean and legume sees should be included in the diet, they must soaked for 48 hours before processing. The soaking will stimulate an enzyme present in the soybean/seed to breakdown the oligosaccharides into simple sugars. Seeds utilize these sugars as energy source for germination.

While carbohydrates are not the main source of energy for Koi, if present in diet, they do spare protein for tissue building activity. Also, carbohydrates in excess are converted into fats for storage.

Vitamins
Water soluble vitamins like Vitamin B and Vitamin C, if fed in excess, will be excreted in urine because they are water soluble. Where as Fat soluble vitamins like Vitamin A, D, E and K  will be stored in tissues if consumed in excess, because they are not soluble in blood and hence can not be transported to form urine. The excess in tissues/organs cause toxicity.

Vitamin B has several members in its family but only 8 have been studied in fish nutrition. These are Thiamine, Riboflavin, Pyridoxine, Pantothenic Acid, Niacin, Biotin, Folic Acid and Cyanocobalamin.

In addition to Vitamin B & C, fish need additional water soluble nutritional factors like Choline, Inositol, para-Aminobenzoic Acid, Lipoic Acid and Citrin

Thiamine or B₁ :Requirement is 2-3 mg/kg dry diet. Thiamine is essential for good appetite, normal digestion, growth, fertility and nerve function. Common sources of Thiamine are dried peas, beans, cereal bran, dried yeast. Moisture, exposure to light and air can inactivate Thiamine. Clams, shrimp, mussels, beans and mustard seeds have an enzyme that breaks down thiamine. These substances can be added to diet only after steaming or drying so the enzyme is destroyed. Deficiency can cause poor appetite, muscle loss, convulsions, instability or loss of equilibrium, swelling, poor growth

Riboflavin or B₂: Requirement is 7 - 10 mg/kg dry diet. Riboflavin is very important in vision, light adpatation  of the eye. Riboflavin is present in milk, germinated grains, peanuts, soybeans, eggs and organs like liver, kidney and heart. exposure to light converts Riboflavin into inactive for Lumiflavin. Deficiency results in cataracts, cloudy eye, photophobia, poor appetite, dark pigmentation.

Niacin or B₃: Requirement is 30 - 50 mg/kg dry diet. Active form of Niacin is Nicotinic Acid amide or Niacin Amide. This is an important factor in energy production since nearly all metabolic oxidation/reduction reactions depend on this vitamin. Fish lose appetite and exhibit jerky or difficult motion and weakness. Muscle spasms can be seen even when fish are resting. Niacin deficient fish are prone to sun burns and turn dark. and show skin hemorrhages. In later stages, fish develop swollen abdomen and poor growth.

Pantothenic Acid or B₅: Requirement is 30- 40 mg/kg dry diet. This vitamin is involved in fat and carbohydrate metabolism, synthesis of proteins, steroid hormones, antibodies and neurotransmitters. Fish deficient in pantothenic acid develop clubbed gills, become prostrate or sluggish, lose appetite, exhibit cell deaths leaving scars. yeast, fishmeal and organs like liver, kidney and heart are great sources of pantothenic acid. Fish pellets contain calcium salt of pantothenic acid which is generally stable but high heat used during processing can destroy the vitamin. The form present in cereal bran is unavailable to fish and therefore it is not a good source.

Pyridoxine or B₆: Requirement is 5 - 10 mg/kg dry diet. Pyridoxine is involved in fat and protein metabolism, production of vitamin B₃ and synthesis of hemoglobin and neurotransmitters. In fry and juvenile stages, carp are carnivorous and feed on large amounts of animal protein (50-60% of their diet). So they need Pyridoxine in good amounts to digest the protein consumed. Rapid exhaustion of Pyridoxine can result in death of entire population in as little as 3 weeks! Deficient diet can result in nervous disorders, gasping. flexing gill operculum to breathe, and bloated abdomen (abdomen may have colorless fluid). They can also develop rigor mortis a few hours before dying.

Yeast, egg yolks, cereals, and organs like liver and glands are good sources of Pyridoxine. Some forms of Pyridoxine is destroyed on exposure to light, air and moisture but plant based forms are generally stable.

Biotin or B₇: Requirement is 1-15 mg/kg dry diet. Biotin plays an important role in the metabolism of lipids, proteins and carbohydrates. Yeast, milk products, egg, nut meats and organs like liver and kidney contain Biotin in significant amounts, But, Avidin, a protein found in egg whites, binds biotin and makes  it unavailable to fish and other animals. Cooking eggs destroys Avidin and releases Biotin. Biotin is one of the expensive vitamins to add in the diet. Cheap foods may not have enough Biotin supplement. Loss of appetite, color and muscle mass,  poor growth and fractured red blood cells are a few signs of Biotin deficiency.

Folic Acid or B₉: Folic acid is essential for cell division, production of blood cells and has a role in production and hatchability of eggs, growth of fry and juvenile fish and blood glucose regulation. Folic acid is destroyed on extended storage or on exposure to sunlight. Anorexia, anemia, poor growth, lethargy, fragile fins and dark skin pigmentation are the main symptoms of Folic acid deficiency. Yeast, green vegetables, organs like liver, kidney and glands, fishmeal are good sources of folic acid. Insects contain a yellow pigment called xanthopterin which has folic acid activity, though not as strong as Folic acid itself. In a pond, insects and algae can provide Folic acid to some extent. Since Folic acid is destoyed easily, one must make sure to give supplements because Folic acid helps in fighting diseases.

Cyanocobalamin or B₁₂: Cyanocobalamin, like Folic Acid is essential for production of blood cells, proteins and nerve sheaths. It is stored in fish tissues and is exhausted after 12-16 weeks. Poor appetite, low hemoglobin, fractured blood cells resulting in anemia are some of the symptoms that point to Vitamin B₁₂ deficiency. Rich sources of vitamin B₁₂ are fish meal and organs like liver, kidney, and glands. But tissues must be carefully processed since cyanocobalamin is easily destroyed by light, heating or in acidic solutions and on storage.

Choline:Requirement is 500 -600 mg/kg dry diet. A derivative of Choline, acetylcholine, is a neurotransmitter. It is involved in fat metabolism and prevents development of fatty liver. Poor growth despite eating good feed is a sign that fat metabolism is affected. Bleeding kidney and gut are also symptoms of choline deficiency. Rich sources are wheatgerm, beans and organs like brain and heart. If choline is in direct contact with Vitamin E and K it can inactivate them. if you are supplementing vitamins, make sure to buy gelatin coated tablets.

Myo-Inositol: Requirement is 200 -300 mg/kg dry diet. Also called muscle sugar, Myo-inositol is involved in fat metabolism. Fish can not pass food through fast enough in their gut when Myo-inositol is deficient in their diet. They develop bloated bellies and show poor growth as a result.  Myo-inositol is present in large amounts in wheat germ, dried peas, beans and organs like brain, heart and glands, citrus fruits and dried yeast. Micro organisms present in the gut can synthesize some amounts of Inositol. If plant fiber is present in the feed, it will bind to Inositol making it unavailable for fish.

para-Amino Benzoic Acid: It is a stable growth promoting vitamin. Micro organisms in the gut need this factor to synthesize Folic Acid (Vitamin B₉). No abnormal indication in growth appetite, mortality if deficient in the diet.

Lipoic Acid: This vitamin is soluble in both water and fat.  Glad tissues are good source of Lipoic acid.

Vitamin C or Ascorbic Acid: Requirement is 30 -50 mg/kg dry diet under normal circumstances but may need up to 1gm in stress situations and for wound repair. Also small fish need more Vitamin C compared to adults because large Carp can synthesize Vitamin C to some extent. Vitamin C helps in fighting diseases by stimulating the immune system. It also helps in detoxifying drugs, production of steroidal hormones, in making collagen for skin and bone formation and in maturation of blood cells. Vitamin C is very stable in acidic conditions but is highly unstable in solutions above pH 7.0, and gets oxidized on exposure to air in presence of metals like copper, iron. It is destroyed in heat as well. Loose teeth, poor bone formation, anemia have been reported in fish with Vitamin C deficiency. Overgrown jaw and snout, bleeding at the base of fins were common symptoms seen in all fishes. If continued, the deficiency results in malformation of spine (bent fish) and the damage is irreversible. Supplementing Vitamin C is quite easy since it is present in all citrus fruits, cabbage, fresh insects and organs like liver and kidney.

Fat soluble vitamins, when fed in excess, can not be excreted because they are not soluble in blood. Instead they accumulate in tissues resulting in toxicity. Symptoms of toxicity for Vitamin A and D are the same as deficiency symptoms of those vitamins. So, often when symptoms are noticed, fish are given more vitamin D and A, making the problems worse. Another problem is the addition of fish oil to fish food in order to add calories cheaply. Fish oil is rich in vitamin A and D.

Vitamin A or Retinol: Requirement is 1000 - 2000 I.U/kg dry diet. Vitamin A is abundant in freshwater fish and vitamin A₁ is found in saltwater fish. Vitamin A is important for night vision resistance to infection and skin maintenance. Too much vitamin A causes enlargement of liver and spleen, poor vision, bleeding at the base of fins and abnormal bones and fused spine. Fish need vitamin A for growth in light but not in dark. If fish are grown in very low light levels and have low stress and no infections, they can do away with very low levels of vitamin A. Whales avoid hypervitaminosis A by converting the excess vitamin A into an inactive form called kitol and store it in their liver. Kitol becomes active only upon heating it to 200*C. Cod liver oil is the best source of Vitamin A. Fish food manufacturers routinely add fish oil to the feed to reduce the amount of expensive fishmeal or carotenes. This can quickly lead to hypervitaminosis A. Feeding color enhancing feed containing large amounts of carotenes can also lead to hypervitaminosis A.

Vitamin D or Cholecalciferol: Requirement is 2-3 mg/kg dry diet. Vitamin D is necessary to balance the amounts of calcium and phosphate in blood. It promotes the absorption of calcium in the gut. but since fish can extract calcium present in water through the gill membrane, they may not use Vitamin D for that purpose. Vitamin D deficiency/hypervitaminosis hasn’t been studied in detail. But fish oil additives in the feed can raise Vitamin D levels more than required. For example, tuna liver oil has 1000 times the requirement of Koi.

Vitamin E or Tocopherol: Requirement is 80 - 100 mg/kg dry diet. Tocopherols are stable to heat and acids in the absence of oxygen but are highly unstable in the presence of oxygen. This is why they are good anti-oxidants. They protect vitamins, unsaturated fatty acids from oxidizing by reacting with oxygen themselves. They are also sensitive to light. Bulged eyes, distended belly, anemia, poor growth are a few signs of deficiency of Vitamin E. Deficiency of Vitamin E can result in deficiency of other vitamins and fatty acids because they can get oxidized in the absence of vitamin E. If diet contains large quantities of poly unsaturated fatty acids, large amounts of Vitamin E must be supplied.

Vitamin K: It is involved in the synthesis of blood clotting factors and therefore is responsible to maintain a normal rate of blood clotting. For fish, especially, the fast clotting is important because they live in water. Deficiency can result in bleeding in gills, eyes and wounds. Green leafy vegetables like alfalfa leaves are the best source of vitamin K. Spoiled alfalfa leaves may contain an antagonist, Dicumarol. Dicumarol is used as an anticoagulant (prevents blood clotting) for people with heart diseases. Warfarin, a type of rat poison, is also an antagonist of Vitamin k and is used as a drug for people with heart diseases. Vitamin K can also be lost on storage or in moisture.

Minerals
Fish need calcium, phosphorous, sodium, molybdenum, chlorine, magnesium, iron, selenium, iodine, manganese, copper, cobalt and zinc. Minerals are constituents of rigid structural body parts like bones, teeth, nails. They are also involved in maintaining the body pH, preventing fish from imploding or exploding by maintaining ionic balance with the water. Some of them are components of pigments, enzymes, hormones.

Calcium and Phosphorus: The metabolism of these minerals are intertwined. They are combines with each other in bones and deficiency of one results in deficiency of both. About 99% of calcium and 80% of phosphorus in fish are present in bones, teeth and scales! The ratio of calcium to phosphorus is roughly 2: 1. The remaining 1% of calcium is present in organs and tissues and in body fluids. Some of this 1% calcium is bound to proteins and is not movable. Other part is in phosphate or bicarbonate form and is movable. This part is responsible for calcium and phosphorus nutrition. The part that is in blood and fluids, maintains ionic balance and muscle activity.

Koi can extract calcium from water through their gills but not phosphorus. In fact, even though plants contain phosphorus in the form of phosphates, they are bound to plant fiber and Koi can not absorb them. We must supply phosphorus in the diet. Phosphorus is absorbed better as water temperature rises and if glucose is present in the food. Since the level of phosphorus determines the level of calcium, it is important to supply enough phosphorus in the diet. Calcium to phosphorus ratio in Koi body is 1: 4 and 1: 2 in Koi skeleton. Optimum concentration of Phosphorus in the diet must be 0.7% or 7g per kg dry diet whereas 5g per kg dry diet of calcium is sufficient..

If phosphorus is lacking or low in the diet, fish develop deformed backs and heads. Fishmeal is rich in both calcium and phosphorus as long as it does not contain bones. Koi can not digest phosphorus from bones. Phosphorus availability for grains is 33%, for soybean meal is 40% and for fish meal is about 50%

Magnesium: About 60% of magnesium in carp is stored in the skeleton. The remaining 40% is in organs, muscle tissues and body fluids. Magnesium is part of many enzymes and helps in metabolism of fats, sugars and proteins. Like calcium, fish can extract magnesium from water if present in high concentration. But usually, concentration of magnesium is low in freshwater. Koi need 500mg per kg dry diet of Magnesium for normal growth. Magnesium deficient diet can result in poor appetite, sluggishness, spasms and convulsions.

Sodium helps Koi in maintaining pH of their blood. Maintaining pH is very important because most enzymes work efficiently only in a small range of pH. Sodium also helps in ionic balance along with chlorine and potassium. Optimum levels of sodium in the diet should be between 103gm per kg dry diet.

Potassium in involved in nerve function and ionic balance. Like sodium, 1-3g/kg dry diet of potassium must be supplied in the diet.

Sulphur is part of amino acids and collagen. it also helps in detoxification of drugs and poisons. About 3-5g/kg dry diet of sulphur must be provided.

Chlorine occurs as chloride (part of salt or sodium chloride) in blood and other body fluids and is involved in ionic balance. Chlorine is supplied as salt up to 1-5g/kg dry diet

Iron is important in production of blood cells, respiratory enzymes and also involved in energy production. About 50-100mg/kg dry diet must be provided in the diet because deficiency results in anemia.

Copper is part of blood cells and some enzymes. Koi need 1-4g/kg dry diet for normal function.

Manganese is part of many enzymes and is involved in bone formation and blood cell regeneration. About 20-50mg per kg dry diet is sufficient as supplement.

Cobalt is the metal component of Vitamin B_12. 5-10mg per kg dry diet of Cobalt must be given in order to maintain blood clotting at normal levels.

Iodine is necessary to regulate oxygen use. Its deficiency produces goiter-like conditions. 100- 300mg per kg dry diet is necessary.

Zinc is essential for insulin and enzymes that maintain ionic balance. Zinc deficiency causes cataracts, erosion of fins and skin. Since zinc also helps fighting diseases it is important to supply 30-100mg per kg dry diet in the food.

Koi also need molybdenum, chromium and fluorine in trace amounts.

Ash or Fiber: Fiber is not digestible by fish. They are added as fillers and to make the food move easily in the gut.

Bibliography:
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Wilson, 1989

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