Biochemistry, Ammonia

Article Author:
Shamim Mohiuddin
Article Editor:
Divya Khattar
Updated:
4/29/2019 7:35:29 PM
PubMed Link:
Biochemistry, Ammonia

Introduction

Ammonia production occurs in all tissues of the body during the metabolism of a variety of compounds. Ammonia is produced by the metabolism of amino acids and other compounds which contain nitrogen. Ammonia exists as ammonium ion (NH4+) at the physiological pH and is produced in our body mainly by the process of transamination followed by deamination, from biogenic amines, from amino groups of nitrogenous base like purine and pyrimidine and in the intestine by intestinal bacterial flora through the action of urease on urea. Ammonia disposal takes place primarily by the hepatic formation of urea. The blood level of ammonia must remain very low because even slightly elevated concentrations (hyperammonemia) are toxic to the central nervous system (CNS). A metabolic mechanism exists by which nitrogen is moved from peripheral tissues to the liver for its ultimate disposal as urea, while at the same time maintaining low levels of circulating ammonia.

Fundamentals

The amino acids take part in certain common reactions like transamination followed by deamination for production of ammonia. The amino group of amino acids is utilized for formation of urea which is an excretory product for protein metabolism. The amino acid is transaminated to produce a molecule of glutamate. Glutamate is the one amino acid that undergoes oxidative deamination to liberate free ammonia for the synthesis of urea. Once free ammonia is formed in peripheral tissues, it must be transferred to the liver for the conversation to urea. This is carried out by the ‘glucose-alanine cycle”. In the glucose-alanine cycle, alanine which is formed by the transamination of pyruvate gets transported in the blood to the liver, where it is transaminated by alanine transaminase to pyruvate. The non-toxic storage and transport form of ammonia in the liver is glutamine. Ammonia is loaded via glutamine synthetase by the reaction, NH3 + glutamate → glutamine. It occurs in nearly all tissues of the body. Ammonia is unloaded via glutaminase by a reaction, glutamine --> NH3 + glutamate. It specifically occurs in kidneys and intestine and in very low concentration in the liver. This reaction is induced by acidosis.

Cellular

In nature, ammonia exists as both NH3 and in the ionic form of ammonia as ammonium ion (NH4+). A buffering reaction: NH3+ H+ --> NH4+ is used to maintain the relative amount of each form. Under biological conditions, the pKa of this reaction is about 9.15 and this reaction occurs almost instantaneously.  As a result, the majority of ammonia under physiological conditions exists as NH4+, and only about 1.7% of total ammonia presents as NH3 at pH 7.4. Ammonia is a very small, uncharged particle. Due to this character of ammonia, it was initially believed that ammonia is highly permeable across the lipid membrane because of the maintenance of proper diffusion equilibrium. But later, after thorough and extensive studies this was refuted. Instead, it was seen that though ammonia is an uncharged particle, the asymmetrical arrangement of positively charged hydrogen ions around a central molecule of nitrogen converts this ammonia molecule into a relative polar particle. Ammonia has a molecular dipole moment of 14.7 D which denotes the degree of separation between the positively and negatively charged particles. In contrast, HCl has a dipole moment is 1.08 and the water molecule has a dipole moment of about 1.85. Due to this charged polarity ammonia has limited and minimal permeability through lipid membranes. This typical character of permeability results in the development of transepithelial gradient of ammonia and this is demonstrated to be present in the kidneys. In the absence of specific transport proteins ammonia also have a restricted property of permeability across lipid bilayers. Due to the inability for the transport of ammonia through the lipid bilayer in the plasma membrane, the hypothesis of transporting of NH4+ by “NH4+ trapping” was introduced, although the accuracy of this concept has not been fully established. Ammonium ion (NH4+) has very poor permeability across the biological membrane in the absence of appropriate transporter. There is no detectable permeability in some tissues such as the apical membrane of collecting duct segments. However, transport of ammonium ion (NH4+) across the biological membrane can occur by specific proteins and is particularly crucial for renal ammonia excretion. Due to the particular biological character of ammonium ion in hydrated form, these proteins can be used to transport this specific ion. Ammonium ion (NH4+) and potassium ion (K+) show almost identical biophysical character when examined in aqueous solutions. This particular unique character allows ammonium ion to be effectively transported at the transport site of potassium ion.[1]

Function

In response to an acid challenge, the production of ammonia and its excretion are major mechanisms by which the kidney produces bicarbonate.[2]Under physiological conditions when the body is exposed to an acid environment, the kidney stimulates the production of ammonia and its excretion. The primary source of ammonia is glutamine which gets excreted in the urine. The proximal tubule is the main site of ammonia formation, and the effective rate of delivery of glutamine in this site not only depends on the sufficient delivery of glutamine but also on the ability of proximal tubule to take up that particular glutamine delivered. The acidotic condition stimulates the delivery as well as augmenting the transport of glutamine into the kidney. SNAT3/Slc38a3 is a glutamate transport protein, and the amount of this increases with an increase in uptake of glutamine and resulting acidosis. Enzymes responsible for the production of ammonia are upregulated by the acidotic condition that leads to augmented production of ammonia from proximal tubules of the kidney. This acidosis also stimulates increased secretion of ammonia into the lumen which then results in increased transport of ammonia towards the thick ascending limb, leading to enhanced absorption and formation of ammonia in medullary interstitium.[3]

Testing

It is clinically relevant to determine the level of ammonium in the urine to determine the capacity of kidneys for an appropriate response to an acid challenge. Kidneys excrete increased amounts of ammonia in acidotic conditions than normal acid-base balance conditions.  There are several methods for estimation of ammonia excretion through the kidney. One of the most appropriate and widely accepted methods is to measure the urinary anion gap and urinary osmolal gap.  Urinary anion gap is determined as UNa+ + UK+ −UCl-. This particular method is beneficial based on the assumption that urinary ammonium ion is excreted only in association with the chloride ion. But this method is not useful for other ions like sodium, potassium, or glucose and urea nitrogen. For this, urinary osmolal gap estimation is necessary. The urinary osmolal gap is determined by Uosm−[2×(UNa++UK+)+UUN/2.8+Uglucose/18)]Uosm−[2×(U+U)+U/2.8+Uglucose/18)]. One can assume that in the absence of any osmotically active material like mannitol or unmeasured cations, the urinary osmolal gap only shows the ammonium ion concentration with its anion. However, the gold standard of measurement of urinary ammonium ion is the same as the enzymatic assay to measure the blood ammonium ion levels.[4]

Clinical Significance

In chronic kidney diseases (CKD), the kidney is unable to produce and excrete an adequate quantity of ammonia which leads to retention of acid and formation of metabolic acidosis.[5] With the progression of kidney disease, the glomerular filtration rate simultaneously falls and leads to increased production and excretion of ammonia by the remaining functioning nephrons. Subsequently, the remaining functioning nephrons cannot sustain the gradual increase of dietary acid load and lead to excessive retention of acid inside the body.[6] In CKD, the kidney is unable to take in or metabolize glutamine which is the substrate for the production of ammonia. Glutamine uptake and metabolism contributes to only about 35% of ammonia production. The rest comes from other amino acids derived by the breakdown of peptide linkages. Further studies show that glutamine supplementation can increase the formation of ammonia in the normal individual but not in patients of CKD, although serum level of glutamine is high in both cases. This unique phenomenon in case of CKD exists due to the reduction of glutamine transporter SNAT3/Slc38a3.[4] Studies performed in nephrectomized rats show that other defects can be seen in the production of ammonia and transport. Researchers found in the animal model of CKD that despite the urinary acidification, the defect was in the net excretion amount of acid.  It was also seen, in comparison to normal control, the delivery of ammonia shows a marked elevation at the peripheral accessible portion of the proximal renal tubule. Research also observed that ammonia is at a lower concentration in the loop of Henle, which allows for the escape of more ammonia mainly from the cortex of nephron, and it then enters back in the renal vein and returns to the central circulation. This particular property decreases the amount of ammonia in the medullary interstitium which leads to a decreased concentration gradient between medullary interstitium and collecting duct lumen. This specific defect of luminal entrapment of ammonia in the collecting duct is believed to correlate with distal delivery of bicarbonate that leads to increased reabsorption of bicarbonate, reduction of formation of titratable acid and secretion of ammonia. Recent studies shown in polycystic kidney model show that the decrease in ammonia excretion in urine is due to the decrease of ammonia transporter called RhCG. However, this hypothesis has been refuted with the findings achieved in the remnant kidney, which shows that the distribution of RhCG transporter protein increases in apical and basolateral portion. So, in patients of chronic kidney disease, despite the presence of acidosis, the production, as well as excretion of ammonia, are seen to be reduced. Thus normal acid-base balance is disrupted in case of chronic kidney diseases.[4]

Hyperammonemia (elevated ammonia concentration in systemic circulation above the normal range of approximately Greater than or equal to 65 micromoles) correlates with liver failure and other significant causes of toxicity of skeletal muscle. So liver disease associated with hyperammonemia is an apparent cause for muscle wasting disorders. A recent study showed that ammonia lowering therapy in hyperammonemic portocaval anastomosis rat models resulted in improvement of the phenotype of muscle and metabolic activity of the protein. Though it is not very clear what the exact mechanism of myopathy is; the assumption is that detoxification of ammonia takes precedence over protein synthesis in muscles. An elevated level of ammonia has also been proposed to increase muscle breakdown through the activation of autophagy, contributing to the loss of muscle mass associated with cirrhosis. Additionally, alcohol correlates with an elevated level of serum ammonia can exacerbate the muscle protein metabolism impairment and elevate the risk of associated hepatic myopathy. This hypothesis supports the observation that patient suffering from alcoholic liver disease have a higher incidence and degree of muscle wasting than the hepatic disease due to toxic or other infective causes.[7]

Hemorrhagic shock is also known to be a cause of elevated blood ammonia levels. Excessive hemorrhage reduces the total hepatic blood flow which causes ischemia in periportal to the centrilobular area of the liver, and that leads to necrosis in in patients in irreversible shock. The pericentral hepatocyte is responsible for the synthesis of glutamine, and periportal hepatocyte is responsible for urea synthesis. High concentrations are the result because of the decreased capacity of detoxication results due to dysoxia of these cells.[8]


References

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