Importance of Methylcobalamin in Animals
Ramesh Jagtap, D J Kalita and Dr. Laxmikant Gode
Introduction
Cobalamin, commonly called as Vitamin B12 is a water-soluble vitamin, which is vital for cellular growth, proper functioning of the nervous system and for the creation of blood cells. It is dietary essential in most of the animal species as these species unable to synthesize cobalamin. Cobalamin functions as cofactor for several enzyme systems involved in nucleic acid synthesis, fatty acid synthesis, amino acid metabolism, myelin synthesis and haematopoiesis.
There are three natural forms of vitamin B12, methylcobalamin (MeCbl), adenosylcobalamin (AdCbl), and hydroxycobalamin (OHCbl) (Dubeski P.L. 1992). Methylcobalamin is the most bioavailable, neurologically active and best utilized form of vitamin B12. Methyl B12 is more effective than traditional forms of B12 as there is no need for conversion in the body and is better retained by the liver and other tissues. It is an active coenzyme of the vitamin B12 analogue that is essential for cell growth and replication.
Methylcobalamin is an essential part or cofactor in several important enzyme systems which carry out several basic metabolic functions and thus, it is vital in metabolism of carbohydrates, lipids, amino acids and DNA (Herdt and Hoff, 2011; Forrellat et al., 1999). Also, it functions by making a substance called myelin, which covers nerve fibers and protects them.
Role in Nucleic Acid metabolism
Methylcobalamin plays an important for DNA synthesis and ensures structural stability of important regions of the chromosomes such as the centromeres and the subtelomeric DNA. As a methyl-donor, it participates in the monocarbonic acid metabolic pathway and plays a critical role in DNA methylation, which is especially important during embryogenesis and carcinogenesis. DNA methylation is catalyzed by DNA methyl-transferases that transfer methyl groups from S-adenosylmethionine (SAM) to cytosine. Methionine is critical for the methylation of various biological molecules, including DNA (Rizzo and Lagana, 2020). Further, methionine synthetase is crucial for nucleic acid synthesis, as tetrahydrofolate is a precursor of purine and pyrimidine synthesis. Low serum levels of vitamin B12, induce alterations in DNA synthesis and may lead to DNA damage.
Role of cobalamin in functioning Nervous system
Methylcobalamin is essential for the preservation of the myelin sheath around neurons and for the synthesis of neurotransmitters. Cobalamin deficiency can lead to increase in serum homocysteine, which hamper cognitive functions and can accelerate neurodegeneration in CNS as well as spinal cord. Methylcobalamin is the only form of vitamin B12 which can directly participate in homocysteine metabolism. In addition, converting homocysteine to methionine via methyl B12 generates an increased supply of SAMe (S-adenosyl methionine), the body’s most important methyl donor.
Role in Energy metabolism
Cobalamin mainly involved in two enzymes systems as shown in fig. Cobalamin has role in energy metabolism through its involvement in two important enzymatic processes.
- Cobalamin is a cofactor in the form of adenosylcobalamin in the reaction catalyzed by enzyme L-methylmalonyl-CoA-mutase (mutase) in which methylmalonyl-CoA is converted to succinyl-CoA, which subsequently enter into TCA (Tricarboxylic Acid) cycle for the production of energy units in the form of ATP and NADPH. Further, the Succinyl-CoA play vital role in the production of energy from lipids and proteins and is also required for the synthesis of hemoglobin, the oxygen-carrying pigment in red blood cells (Shane, 2000).
- Methylcobalamin as cofactor essential for remethylation reaction of homocysteine to methionine by methionine synthase (MS) as shownn in fig 1. Methionine in the form of S-Adenosylmethionine required for most biological methylation reactions including DNA methylation.
Fig.1: Role of Methylcobalamin in two enzyme systems methyl-malonyl CoA mutase and Methionine synthase
Importance of Methylcobalamin in Ruminants
Ruminants’ cattle, buffalo, sheep, goat, camels, etc., obtain required cobalamin from microbial synthesis, mostly by bacteria and archaea present in the rumen. Cobalamin is not present in the plant origin feedstuffs, which are fed to the ruminants. However, the ration containing adequate cobalt (which is a central atom in cobalamin) is required to supply the cobalamin to ruminant animals, as several strains of microbes in the rumen can able to synthesize it utilizing dietary cobalt. But, conversion of dietary cobalt to Vitamin B12 is very inefficient in ruminants, with 3-15% cobalamin production in sheep and in dairy cows efficiency is 7.5-11%. Usually, higher dietary cobalt can lead to higher cobalamin synthesis. Rumen microbes utilize cobalamin for the breakdown of cellulose.
Besides, dietary cobalt content, microbial cobalamin synthesis can also influence by dry matter intake or roughage content of the ration, higher being increases the cobalamin production. While high concentrate content can reduce the cobalamin synthesis by rumen microbes. Apart from this, the vitamin B12 status is depend on efficiency of absorption and the metabolic demands of the co-enzyme dependent functions.
On the contrary, in preruminant lambs and calves, up to age of 6-8 weeks, require dietary cobalamin, as the rumen is not fully developed and not functional for its synthesis (Stemme et al, 2009).
Energy metabolism in ruminants
Besides role in Nucleic acid metabolism and DNA synthesis, functioning of nervous system, in ruminant’s cobalamin is involved in utilization of propionic acid, which is source of energy in ruminants. Due to its important role in propionic acid metabolism, ruminants require more cobalamin than non-ruminant animals.
In the process of utilization of propionic acid, as shown in fig.2., Methylcobalamin can act as cofactor in the form of 5- deoxy-adenosylcobalamin (Ado-cbl) in the reaction catalyzed by enzyme L-methylmalonyl-CoA-mutase, which convert L-methylmalonyl-CoA to succinyl-CoA, which subsequently enter into TCA (Tricarboxylic Acid) cycle and produce energy to the animals. In ruminants, methylmalonyl-CoA plays a unique regulatory role in gluconeogenesis and fatty acid oxidation. Propionic acid being important energy source and precursor for gluconeogenesis, its conversion to succinyl CoA is critical reaction for glucose homeostasis in ruminants.
Fig.2: Role of Methylcobalamin as deoxy-adenosylcobalamin in propionic acid metabolism in ruminants
Cobalamin Deficiency in Ruminants
Deficiency of Cobalamin is mostly due to the deficiency of Cobalt in the ruminants. Cobalamin deficiency led to coastal disease in sheep or wasting disease, ill-thrift or enzootic marasmus in cattle, which is also called as bush sickness in countries like New zeland. As like other animals in ruminants its deficiency can be associated with conditions like accumulation of methylmalonic acid, megaloblastic anemia and pernicious anemia. Cobalamin deficiency has been reported in many parts of the world. Usually, cobalamin deficiency shows nonspecific symptoms, reduced appetite, rough body coat, retarded growth, wasting of muscles, etc. Further, its deficiency often reduces production efficiency and also led to reproductive disorders.
Cobalamin deficiency due to cobalt deficient diet has been shown to negatively affect the immune function of ewes and calves, leading to increased susceptibility to infection in ewes, with particularly serious consequences for the viability of newborn lambs.
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Cobalamin deficiency in ruminants often lead to reduced energy from glycolysis, however it may increase energy from gluconeogenesis or oxidation of lipids. Periods of Cobalamin deficiency can reduce appetite and hence low feed intake also stimulate the mobilization and oxidation of fatty acids of lipids in the liver to supplement the short fall in energy from glycolysis. But due to cobalamin deficiency, levels of methyl malonyl-CoA build up, which inhibit β-oxidation. Thus, lipids are accumulated in the liver of sheep causing ovine-white liver disease (Kennedy et al., 1994).
Cobalamin deficiency and associated conditions in non-ruminant and companion animals
Unlike ruminants, most of non-ruminant animals including pigs are unable to synthesize vitamin B12 efficiently, so instead of cobalt these animals require adequate supplementation of cobalamin. Deficiency can lead to reduced growth rate and anemia, as cobalamin along with folic acid, iron, and copper are essential for synthesis of erythrocytes (NRC, 1998). In few animal species like pigs, horse, chicken including companion animals’ dog and cat, deficiency of cobalamin can occur due to following major causes:
- Pernicious Anaemia: It’s an autoimmune condition, in which antibodies to intrinsic factor are produced and it is unable to bind to cobalamin. Thus, inability of the animals to absorb Vitamin B12 from terminal ileum, its site of absorption.
- Malabsorption: Gastrointestinal disorders, that can affect the production of IF from parietal cells of the gastric mucosa may be at risk of deficiency. Also, gastrointestinal surgeries or damage to intestine, especially terminal ileum, which can affect its absorption can lead to deficiency of Cobalamin.
- Deficient forage or diet of the animals: Chronic dietary insufficiency of vitamin B12 or Vitamin B12 deficient diet can lead to deficiency disorders in few species of animals like pigs, dogs and cats. Indirect deficiency can occur due to cobalt deficient or insufficient diet or ration of the animals.
- Genetic defects affecting Cobalamin metabolism: In many breeds of dogs, including Beagles, Border Collies, Shar Pei’s, Giant Schnauzers and Australian Shepards, the genetic defects are identified in the receptors which bind to Intrinsic factor and thus affect cobalamin absorption. Cobalamin deficiency can also observe due to congenital defects (Lutz etal., 2013; Erles et al., 2018; Grützner et al., 2010) in receptor genes, which affect intracellular metabolism of cobalamin (Eg. Shar Peis, Giant Schnauzers breeds of dog). Breed related Congenital genetic defects not yet identified in cats.
Gastrointestinal disorders are quite common in pet animals, especially Dogs and Cats. The diseases like GI lymphoma, Inflammatory bowel disease, intestinal dysbiosis, exocrine pancreatic insufficiency, short bowel syndrome, pancreatitis can lead to cobalamin deficiency in pets (Kook et al., 2014: Fyfe et al., 2014). Cobalamin deficiency can also occur where pancreatic secretion of intrinsic factor is affected.
In dogs with congenital deficiency, the clinical signs include weakness, lethargy, poor body condition, weight loss, inability to gain weight, cachexia, anorexia, diarrhea, vomiting, dysphagia, oral ulcerations, and proteinuria. Hematopoietic conditions like nonregenerative anemia, neutropenia is also observed in affected dogs. Symptoms can occur at early age of dogs (6-12 weeks), but clinical cases were identified up to 3-4 years of age.
Role of Methylcobalamin in milk production
The direct role of cobalamin or methylcobalamin in improving quality and yield of milk in dairy animals is highly debated in scientific community. In the context, many investigations are encouraging especially respect to improving cobalamin content in milk and increased milk yield.
The established dietary requirement of cobalt in ruminants is 0.1 to 0.2 mg/kg. But the suggested requirement of Cobalamin is quite high in dairy cows that is 0.34 and 0.68-microgram/kg body weight. The high needs of cobalamin in dairy cows are due to the fact that large quantities of propionate produced in high yielding dairy ruminants and for its metabolism and subsequent energy production through gluconeogenesis, cobalamin act as essential cofactor. Furthermore, vitamin B12 production from cobalt by ruminal microorganisms and the low efficiency of vitamin B12 absorption justifies the high need of cobalamin to dairy animals. There are mixed reports on increased milk production with increased supplementation with Cobalt.
However, it is reported that parenteral and oral supplements of vitamin B12, as well as folic acid increased milk production and modified milk composition in dairy cows, which clearly suggest that rumen microbial synthesis of cobalamin may not be sufficient to optimize animal performance, especially milk production (Girad et al., 2001). In most investigations, when the total Co of the diet (basal plus supplemental) was approximately 1 to 1.3 mg/kg, maximum responses were observed in milk production (Stangl et al., 2000). However, such production responses require large amounts of vitamin B12 and folic acid in the diet, which are necessary to increase their plasma concentrations to levels similar to those observed after parenteral supplementation (Girad et al., 2001).
Dietary supplementation of cobalamin and folic acid in multiparous Holstein cows, 3 wk pre to 8 wk post-partum showed improved lactational performance and metabolism. Further it showed increased plasma glucose and alanine and decreased lipids in the liver (Graulet et al.,2007). Similarly parenteral injections (IM) of cobalamin and folic acid in commercial dairy herds showed the better energy state of the animals, increased BCS and decreased fat and protein in early lactation. Further, reduction in estimated body weight loss coupled with a reduction of the fat:protein ratio without effect on milk yield was observed which suggest that energy partitioning in early lactation (Duplesis et al., 2014).
Another study showed increased milk production up to 12% in multiparous cows with parenteral supplementation (IM) of folic acid and vitamin B12 in early lactation from 3 to 16 wk after calving (Preynat et al., 2009). As like many studies it also showed increase in cobalamin and folic acid concentrations in Milk and plasma. Further, increased milk lactose, protein, and total solids yields was reported in same study. With cobalamin supplementation as the milk lactose content increase, the body glucose flux also tends to increase with a similar quantitative magnitude. It is also reported that supplementation of methylcobalamin, increase methionine utilization for protein synthesis through increased protein turnover when methionine was deficient.
Summary
Although in-feed supplementation of vitamins including cobalamin is established practice, however, whether the supplements fulfill requirement of cobalamin for animals especially young calves and dairy animals is highly debated. Cobalamin deficiency due to insufficient or deficient dietary intake of Cobalt, malabsorption, genetic or congenital factors, or due to other factors, create unfavorable metabolic consequences to the animals. Deficiency can ultimately lead to reduced energy balance, decrease in production and compromised reproductive efficiency.
Cobalamin is proven metabolic stimulant and parenteral use of Methylcobalamin, an active form of cobalamin, effective in alleviating deficiency symptoms. This is well explained due to its essential role as cofactor in several important enzyme systems which catalyze many basic metabolic reactions and thus play vital role in metabolism of carbohydrates, lipids, amino acids and DNA. Dairy animals, especially in early lactation has very high requirement of glucose, as it is converted to lactose. Furthermore, dairy animals use propionic acid as preferential source for gluconeogenesis. Methylcobalamin is essential cofactor in propionic acid metabolism in ruminants, especially conversion of propionate to succinate, which subsequently after breakdown produce energy. Moreover, it is reported that parenteral injections of cobalamin, especially with adequate supplementation of folic acid support high milk yield in dairy animals.
Methylcobalamin is well-known nervine stimulant and useful for prevention of neurodegeneration in both central and peripheral nervous system, due to its critical role in preservation of myelin sheath of neurons and synthesis of neurotransmitters. Further, its role in one carbon group transfer and DNA methylation is important during embryonic development.
Note: References, if required, can be provided on request
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