Introduction
Metabolic disease refers to a disorder that causes a problem in the normal metabolic process (converting food to energy at the cell surface). During the metabolism, thousands of enzymes are involved. Metabolic disorders affect the ability of the cell to perform biochemical vital responses, including transport of protein (amino acids), carbohydrates (sugars and starch) or fat (fatty acids) [1]. Metabolic disorders are usually inherited. Congenital metabolic disorders are a group of complex and heterogeneous genetic abnormalities that cause clinical signs of mutation in genetic codes and subsequently reduce or increase the activity of enzymes involved in metabolic pathways. Due to severe clinical manifestations, metabolic disorders are one of the major causes of infant mortality [2]. Hence, delay in the diagnosis and treatment of these diseases leads to various adverse effects, including moderate to severe nervous disorders, mental retardation and death. A single disorder occurs less frequently, but the likelihood of a metabolic disorder syndrome is 1 in total per 1,500 live births [3]. This means that in the wake of a mutation in the structure of the parent genome, a disruption occurs in the process of metabolic enzymes and is transmitted to the next generation. These diseases do not always have obvious symptoms and may be asymptomatic for days, months and years. Symptoms of the disease usually occur when the body is under stress, such as long-term hunger or febrile illness. For early diagnosis of some metabolic diseases, there is a possibility of birth screening. Often, in many cases, these screenings occur when a family previously has a child with a metabolic disorder or in a population with a large number of cases, such as the Tay-Sachs disease in the Ashkenazi Jewish population [4].
Classification of metabolic diseases has many types. The earliest grouping of these diseases is based on the general form of the involved metabolism, and in this grouping, a disorder can be grouped simultaneously into several groups. This grouping can be divided into other groups based on similarity in pathogenesis or disease symptoms.
In the other grouping, these disorders are divided into two main groups; the first group is due to the accumulation of toxic substances due to the blocking of metabolic pathways, including acute or progressive disorders of poisoning or encephalopathy, and the second group, which is accompanied by energy-related disorders. But according to one of the most important categorizations, inherited metabolic disorders are divided into three main groups [5]:
 
1) Intoxication Type: This group of diseases includes disorders in intermediate metabolic pathways that cause accumulation of toxic substances (which are already blocked by enzymes) and lead to acute or progressive intoxication through the accumulation of these compounds, including amino acid refluxes, often organic compounds , Urea cycle disorders, some carbohydrate disorders such as galactosemia and hereditary intolerance to fructose, metabolic disorders of heavy metals, porphyry, and defects in the synthesis or catabolism of neurotransmitters. The symptoms of this group are similar and from the beginning they appear with an asymptomatic course and then with signs of equality including vomiting, seizure, liver failure and so on. Diagnosis of these diseases is possible by chromatography of amino acids in the plasma and urine and urinary organics. The mainstay of their predominant treatment is regimen therapy, and most of these diseases can be cured if treatment be administered in a timely manner [6]. In acute cases, it is necessary to prescribe carnitine, benzoate and vitamins for the elimination of toxic metabolites.
 
2) Type of energy deficiency: According to physiopathology, diseases are classified into cytoplasmic and mitochondrial types. Symptoms of this group of metabolic diseases are due to inadequate energy production or the inability to use energy due to defects in the liver, myocardium, muscle, or brain. Major disorders of the cytoplasmic group include glycogenolysis disorders, glycogenogenesis disorders, creatine metabolism disorder, pentose dysfunction and hyperinsulinism, and hypoglycemia is common symptom [7]. In the mitochondrial group, which includes the deficiency of pyruvate carboxylase, pyruvate dehydrogenase, fatty acid oxidation deficiency, Krebs cycle disorder and mitochondrial respiratory tract, hyper lactic accidemia is the dominant symptom. These patients are usually referred to with impaired growth, severe hypoglycemia, lactic acidemia, hypotonia, myopathy, heart failure, arrhythmia, dysmorphism, congenital malformations, cardiovascular collapse, and sudden death [8]. Treatment in a hypoglycemic group is usually possible, but for patients with high levels of lactate, there is often no treatment except fatty acid oxidation defects.

Table 1) Some symptoms of metabolic disorders

 

3) Complex molecular disorders: This group exhibits a defect in the synthesis or catabolism of complex molecules. Their symptoms are mostly permanent, progressive, and unlike the previous two groups, they are unaffected by the physical condition and type of nutrition, and do not require emergency measures. These diseases usually occur at an older age, except for paroxysmal diseases, as well as a number of lysosomal diseases that are present in their neonatal period [9]. Lysosomal abnormalities, paroxymal disorders, congenital defects of cholesterol synthesis, internal dysfunctional disorders, such as congenital defects of glycosylation of molecules or CDG, and deficiency of alpha-1-antipsychotics are in this category.
Hundreds of hereditary metabolic abnormalities have been identified and become known. Some of the most common and important metabolic disorders are phenylketonuria (Phenylalanine hydroxylase deficiency), Maple syrup disease (Branched-chain alpha-keto acid dehydrogenase complex deficiency), Niemann-Pick disease (Lysosomal enzyme acidsphingomyelinase deficiency), Tay-Sachs disease (With defects of Beta-N-acetylhexosaminid), etc. [2, 5, 7-9]. Symptoms of metabolic diseases, depending on the type of disorder, are widely varying and dependent on nutrition, drug use, dehydration and various diseases. Symptoms may appear suddenly or progressively, but often develop within weeks of birth, and may last for many years [2]. Some of these symptoms are summarized in Table 1 [10].
There are limited therapies for metabolic disorders. The treatment of genetic disorders that is the cause of these diseases is also not possible with today's technology. In addition, many children and adults with metabolic diseases require extensive and rapid medical assistance and in some cases it is necessary to hospitalize these patients [11, 12]. Therefore, a case study of early detection, preventive measures and treatment of symptoms of metabolic diseases is preferable to the treatment of their definitive treatment and is based on the following principles [6]:
- Decrease or remove any food that is not metabolized to the patient's body properly.
-Replacing an enzyme or any other chemical that is problematic, and this is done to make the patient's metabolism as normal as possible.
- Removing those toxic products that have accumulated in the body due to metabolic abnormalities.
- Follow the diet.
- Blood with specific chemicals to remove toxic metabolic products.
The overall goal of birth screening is to detect congenital severe abnormalities in healthy neonates [13]. Robert Guthrie was the first person to diagnose phenylketonuria (PKU) at birth by screening of infants in the Guthrie experiment in 1960, and thus became "the father of screening newborns" (Fig. 1).

 

Figure 1) Robert Guthrie, father of neonatal screening

In general, it can be said that biomarkers are molecules that are present in body fluids and their measurement can be used to evaluate normal biological processes, disease state, or response to treatment. In this framework, early markers for neonatal screening are metabolites that are associated with a modified biochemical pathway (reduced or enhanced) in a particular disease and are diagnostically valuable. However, secondary biomarkers are biomarkers indicative of an increased risk of a metabolic disorder that in this case the ratio between analytes can be mentioned. The biomarkers' rating facilitates the interpretation of the profile of these metabolites and increases the sensitivity of the tests. For example, in renal liver tyrosinemia (type I), increase in the succinic stone is considered as primary biomarker, while tyrosine is secondary biomarker because its concentration may be elevated or normal. In addition, tyrosine is not always a sign of this disease. Tyrosine is also increased in preterm infants or low birth weight infants, and liver function is still not fully developed. The use of an analyte ratio is necessary to properly interpret the metabolite profile. Using these ratios reduces the amount of false positive results to less than 0.01% and improves the effectiveness of screening tests. Since the false-positive outcome causes a lot of stress for families, accurate diagnosis of the disease is of particular importance. According to this description, the purpose of writing this research is to introduce the importance of examining these metabolic disorders at birth.
In this research, Elsivier, Pubmed and Scholar google sites were searched for related Persian and English research. Finally, 38 articles were received, translated, and deeply studied, and the history of neonatal metabolic diseases, clinical manifestations in children and adults, as well as recent advances in the field of early diagnosis of these diseases were examined.
Performing an immunoassay for TSH and thyroxin hormones (which are related to the diagnosis of congenital hypothyroidism) in the 1970s led to adding these two tests to the NBS screening program [13]. Potentially, many tests are suggested to be added to the birth-based screening program. However, of course, it is not possible to complete all these tests during the screening program. The World Health Organization (WHO) has defined a golden standard for disorders that should be addressed in this program. The criteria that the World Health Organization has taken into consideration include the spread of technology on diseases, treatment disorders, and the scientific validity of the tests and the cost of the test. According to the WHO criteria, diseases that are a serious health problem should be included in the birth screening program; history of the illness should be adequately screened; the disease should be identified early; and useful tests to diagnose the disease should be available; diagnostic tests for the disease should be acceptable in the community, and the repeat intervals of the test should be defined. There is a need for effective treatment or action, and diagnostic and therapeutic equipment should be available. There should be acceptable therapy policy and treatment costs should be in line with the benefits of it [14, 15].
Screening for phenylketonuria (PKU) was conducted based on a program that examined the relationship between treatment and dietary intake and lack of mental retardation. By the end of 1960, World Health Organization criteria were defined on the basis of the Wilson and Johnger criteria, which had attempted to implement NBS programs [16]. Screening of phenylketonuria (PKU) met these criteria well. Then, programs were successfully developed in other areas, such as screening for hypothyroidism. However, the increase in the number of diseases that could be detected by multiple tests did not meet all the Wilson-Jiang criteria. Since 2000, double-sided mass spectrometry (MS / MS) increased the number of tests that allowed them to detect a large number of diseases. Therefore, today, the national screening program has begun to develop on a large scale in the United States [17]. In 2015, the American National College of Genetic Medicine (ACMG) has proposed 29 diseases that are more essential for screening birth. The American National College of Genetics offers a definition of screening and effective treatment, along with adequate knowledge of the history of the disease [18].
Birth control screening (NBS) is not the only program to detect a concentration above or below the normal levels of blood metabolites, but ideally, this system combines five parts i.e. screen of newborns, observing abnormal screening, diagnostic tests and confirmation using Specialized laboratory tests, diagnosis and treatment, lifelong monitoring of patients, regular and timed communication between the child, the screening officer, health authorities and pediatricians [19]. Today, different screening recommendations are being made. For example, the screening interval until blood sampling in the United States should be within 24 hours, in Germany for 48 to 72 hours, and in the UK for up to 120 hours. Follow up of two-way mass spectrometric screening reports over a 6-year period of children showed that these controls have reduced mortality rates as well as clinical disturbances. This strategy includes diagnosis and treatment before the onset of symptoms. Economically, every Euro spent in this screening program will save you Euro 25 in social and health costs. The results of many studies indicate that the rate of discriminatory clinical disorders and mortality and the improvement in the health status of the patient are the results of screening [19, 20].
The screening of infants as part of child health care is growing significantly in the world. Many countries monitor phenylketonuria and hypothyroidism in infants, while in more developed countries, screening programs include 20 or more disorders. In other countries of the world, screening for birth is also expanding. Some countries have started screening in recent years.
Many developed countries have expanded their screening program through bi-lateral mass spectrometry technology. However, the screening process is very diverse. For example, screening in Australia is done through two-way mass spectrometry techniques. In the United States, nearly 3 million and 400,000 infants are monitored for over 30 metabolic diseases, and in the Netherlands, 17 metabolic diseases are monitored. In Denmark, the program includes 13 diseases, and in Germany, screening of infants is carried out for 12 diseases, and in the UK they are screening infants only for phenylketonuria and MCAD (medium-chain acyl-CoA dehyrogenase deficiency) [21, 22].
The Middle East and North Africa region (MENA) comprises 21 countries and a population of over 400 million people, with fertility rates of over 10 million per year. This population has a high incidence (25-70%) of consanguinity and includes a high percentage of marriage among the second generation. There are studies that show a high percentage of genetic defects in these countries. This highlights the need for a birth-based screening program. Because of the large number of autosomal recessive genetic abnormalities, it is no wonder that in the Middle East and North Africa region, which have a high rate of family marriage, there is a high prevalence of these diseases. Even in developed countries where the percentage of kinship is less than 1%, screening policies are expanding. This is while, in developing countries, in which many parents have kinship relationship, inherited genetic disorders have a higher rate. Health care is still in its early stages. Less than half of the countries in the Middle East and North Africa have a neonatal screening program that covers only hypothyroidism screening. Lebanon and Saudi Arabia use double-sided mass spectrometry systems for birth screening in terms of metabolic diseases. In 2004, Qatar sent samples to the University of Heidelberg in Germany to screen their babies for metabolic diseases [23, 24].
Neonatal screening results using bilateral mass spectrometry show that the incidence of genetic disorders is quite different in different countries [25]. The high prevalence of metabolic diseases in Qatar, which is associated with high consanguinity, is significant. For example, homocystinuria is in the range of 1 to 1800 in Qatar, which is comparable to 1 to 200 thousand in the United States [26, 27], 1 to 230,750 in Australia and 1 to 316243 in Portugal [28]). The incidence of MCAD is 1 to 4000 in Qatar, while the proportion of the disorder in the European population is 1 to 10,000 to 20,000 [29].
Iran is a large country with many ethnicities living in northern, southern and central provinces, with more than 70 million inhabitants and a birth rate of 1.4 million infants per year. In Iran, screening for infants is mandatory only for hypothyroidism, which accounts for approximately 84% of all newborns in the country. Hypothyroidism is of the main candidates in the screening list of infants since many disruptions are prevalent among populations, screening methods are available and economically viable. [30] In the research on the incidence of hypothyroidism in Isfahan, 1 in 748 live births have been occurred between the years 2005 and 2005, which is approximately 5 to 6 times the global prevalence of the disease. According to this study, the incidence of hypothyroidism in Isfahan is 2 to 3 times more prevalent in Greece (1 to 1800), Saudi Arabia (1 to 1,400), and Turkey (1 to 2354) [31, 32].
In Iran, only limited local programs are being conducted to measure phenylketonuria and glucose-6-phosphate dehydrogenase. The largest study to assess the prevalence of phenylketonuria has been reported by Habib et al. in Fars province, which reported an outbreak of 1.6 per 10,000 people. This result is one of the highest amount ​​reported in the world [33]. Another study was conducted by Farhud and Kabiri on 8633 neonates in Tehran and the result was 1.1 in every 10,000 infants [34].
At present, extensive advances in the laboratory area have led to the identification of infants with metabolic defects, using mass spectrometry techniques, a two-way mass spectrometry [34] (Fig. 2).

Figure 2) Basis and components of the Tandem Mass Spectrometry

In this technique, with this advancement, these diseases can be diagnosed with a high sensitivity with a droplet of dried blood on paper. Mass spectrometry is, in fact, a very powerful technique for detecting a wide range of diseases using a sample and on the basis of the separation of ions based on the ratio of mass to the charge and the measurement of its severity. A mass spectrometric device has 5 components, including an ion source, inlet system, mass analyzer 1, mass analyzer 2, and the detector. First, a sample is introduced to the device and is charged after ionization. These molecular ions reach MS1 and are separated according to the charge / mass ratio [34] (Fig. 2).
These molecular ions then enter the enclosure, where they collide with a neutral gas and become separated. Eventually they are separated by MS2 and then detected. The results are ultimately presented as a mass spectrum based on the mass-to-charge ratio. Two-dimensional mass spectrometry can be used as a very fast technique for identifying and recognizing a large number of congenital metabolic diseases that are not recognized by other techniques, including MCAD deficiency disorder (MCAD). The technology of tandem of copper has many advantages over other existing techniques, which has led to a very wide spread worldwide screening of infants. Some of these benefits include the following [35]:
1) Analysis can be done with very little blood sample or other fluids from the infant.
2) The use of two successive spectrometers that in the first spectrometer, the isolation of the compounds from the initial mixture of the sample is carried out and thereby elimination or minimization of the separation is done by chromatography.
3) An analysis requires a short time (about 2 to 3 minutes).
4) All stages of the test are carried out automatically and around 600 samples can be performed within 24 hours.
Two-dimensional mass spectrometry applications for diagnosis of neonatal metabolic diseases are currently measured by two analyte groups, including aminoacid and acylcarnitine, to detect amino acidopathy, organic aciduria, and oxidation deficiency of acidic fat. Metabolic disorders that can be identified by this technique can be divided into four groups of amino acid abnormalities, urea cycle abnormalities, organic anomalies of acids and fatty acid oxidation abnormalities.
 
Conclusion
Neonatal metabolic diseases are one of the most complex genetic diseases, that due to the wide range of diseases and conflicts that inflict on the baby in many ways, cause complicated symptoms through the involvement of various organs of the body, which will make it difficult to diagnose well. Treatments that are considered for these diseases are not definitive treatments. There is only preservative treatment that will prevent disease progression as far as possible. The best way to prevent the progression of metabolic diseases is to diagnose the disease as early as possible to prevent damage to various parts of the body. Diagnosis of these diseases has been done from the beginning to the present by various methods, but today, the best method and reference method for the diagnosis of a wide range of metabolic diseases is the use of mass spectrometry (MS), which has a high sensitivity to the diagnosis of neonatal metabolic diseases.

Acknowledgements: The case was not found by the authors.
Ethical permissions: The case was not found by the authors.
Conflict of interests: The case was not found by the authors.
Financial support: This study was supported by Sarem Fertility and Infertility Research Center and Sarem Cell Research Center.
Contribution of authors: Sara Samari (First author), author of the article/main author/author of discussion (%20); Behnam Younesi (Second author), author of the article/helper author (%20);Peyman Lashgari (Third author), author of the article/helper author (%20); Majid Asiabanha Rezai (Fourth author), author of the article/methodology/main author/author of discussion (%40).