Neermala Ragoonanan

BIOL 2360 – Biochemistry IIA


What is metabolomics?

Metabolomics combines strategies to identify and quantify cellular metabolites using analytical methods such as gas chromatography, liquid chromatography and nuclear magnetic resonance spectroscopy. These analytical approaches are done to analyze different cell products such as those from gene expression (transcripts), proteins, and metabolites. All of these so-called ’omics approaches including genomics, transcriptomics, proteomics and metabolomics are considered important tools to be applied and utilized to understand the biology of an organism and its response to environmental stimuli or genetic perturbation.

There are four approaches in metabolomics: target analysis, metabolite profiling, metabolomics, and metabolic fingerprinting. Target analysis includes the determination and quantification of a small set of known metabolites (targets) using one particular analytical technique for the compounds of interest. Metabolite profiling aims at the analysis of a larger set of compounds, both identified and unknown with respect to their chemical nature. Metabolomics employs complementary analytical methodologies for example, LC-MS/MS, GC-MS, and/or NMR, in order to determine and quantify as many metabolites as possible, either identified or unknown compounds. Metabolic fingerprinting is where mass profile of the sample of interest is generated and then compared in a large sample population to screen for differences between the samples. When signals that can significantly discriminate between samples are detected, the metabolites are identified and the biological relevance of that compound can be revealed thus reducing the analysis time.

Metabolomics can be used for a large range of applications, including phenotyping of genetically modified plants and substantial equivalence testing, determination of gene function and monitoring responses to biotic and abiotic stress. Metabolomics decreases the gap between genotype and phenotype by providing a more comprehensive view of how cells function, as well as identifying novel or striking changes in specific metabolites. It can provide new hypotheses and new targets for biotechnology.

Metabolic profiling or metabolomicics is referred to as being either targeted or non-targeted:

TARGETED: In the targeted approach, specific metabolites of known identity are profiled. In the case of targeted MS, this involves the addition of multiple stable isotope-labelled standards to the biological sample before the extraction and derivatization steps to control for differences in analyte loss during sample processing and to compensate for ionization-suppression effects. Targeted methods provide an excellent survey of metabolic fuel selection and a profile of energy-yielding metabolic pathways, including elements of mitochondrial metabolism.

NON-TARGETED: In non-targeted profiling it involves the use of NMR or MS for simultaneous measurement of as many metabolites as possible in a biological specimen. These approaches are generally used to compare two biological or clinical states and to report on differences between the two states based on peak areas of raw spectral data.


How has metabolomics contributed to the understanding of disease mechanisms and how has it contributed to the improvement or creating strategies for treatment of these diseases?

  • Human diabetes and insulin resistance

Targeted mass spectrometry (MS) based metabolic profiling has been increasingly applied to studies of human conditions. The profiling of an obese person was done in a research conducted that revealed that branch chain fatty acid catabolism correlates with insulin resistance. The results from this research showed that metabolomics can provide a more detailed picture of metabolic status of normal and pre-diabetic subjects, which can be used for further development and could contribute to more exact sub-classification of different forms of diabetes, leading to the development of more effective drugs.

  • Human cardiovascular disease (CVD)

In a study conducted using MS-based metabolic profiling the application of metabolomics was done to determine metabolic lesions in heart failure and myocardial infarction (MI). The growing number of metabolomics studies in the area of heart failure may ultimately facilitate optimal design of perioperative treatment regimens based on the particular form of cardiovascular disease and the metabolic status of the heart. Comprehensive metabolic profiling, or “metabolomics” is increasingly being applied to CVD, leading to recent discoveries with both form and function implications.

Metabolomics profiling of coronary artery disease (CAD): Targeted MS/MS-based methods were used to profile 45 plasma acylcarnitines and 15 amino acids in a larger study of CAD. With the use of principal components analysis for data reduction, two principal components analysis–derived metabolite factors were found to be associated with CAD: one is composed of branched-chain amino acids (BCAAs) and their associated metabolites and one is composed of urea cycle metabolites, including arginine and citrulline. These metabolite clusters discriminated individuals with CAD from those without CAD in both discovery and validation data sets.

Myocardial infraction:  identified certain proteins such as troponin I, troponin T, C-reactive protein, and B-type natriuretic peptide as diagnostic markers for CVD events and heart failure.

Another recent study has identified a fascinating link between the diet, gut microflora, host metabolism, and metabolomic biomarkers of risk for incident CVD events. The study used a non-targeted LC-MS–based metabolomics approach to profile stable patients who subsequently experienced MI, stroke, or death over the ensuing 3-year period compared with age- and sex-matched control subjects who did not experience events.

Integration of Genetics and Metabolomics for the Identification of Novel Disease Pathways:

 A potential avenue for translating metabolomics-derived biomarkers to disease mechanisms is the integration of metabolomics with other “omics” methods. Human genome-wide association studies have mapped loci associated with polygenic disorders like CVD and diabetes mellitus, but they account for only a small fraction of these diseases and have made limited contributions to knowledge-based therapeutic interventions. This results in part because both conditions are actually a family of diseases in which genetic variability, environmental factors and resultant perturbations in metabolic control within multiple tissues and organs combine to disrupt homeostasis and tissue functions.

Additional Readings:

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Brouwer, Ingeborg. 2010. ‘Effect of Animal and Industrial Trans Fatty Acids on HDL and LDL Cholesterol Levels in Humans – A Quantitative Review.’ Accessed April 05, 2013.

Effects of Animal and Industrial Trans Fatty Acids on HDL and LDL Cholesterol Levels in Humans- A Quantitative Review:

          Trans fatty acids can be obtained from industrial hydrogenation of vegetable oil and fish oils (artificial trans fatty acids) or from the biohydrogenation from ruminant animals such as cows and sheep (natural trans fatty acids). The consumption of these hydrogenated products results in the increase or decrease of HDL and LDL lipoproteins in the body which places a risk on a person’s heart.

            In this report 39 studies were conducted using persons with controlled diets. Twenty-nine used industrial trans fatty acids, six used ruminant trans fatty acids while seventeen used conjugated trans linoleic acid (CLA). Linear regression analysis was uses to determine if these individuals were affected. The slope of the line for LDL to HDL ration was steeper for trans industrial fatty acids than for ruminant fatty acids or CLA. Statistical analysis was used to compare trans fatty acids with saturated fatty acids.

            The results indicated that there was significant weight loss and gain for some individuals with an increased risk of heart and liver disease. There is a quantitative comparison of the effect of ruminant trans fatty acids and CLA with industrial trans fatty acids on blood lipoproteins in humans. The analysis shows that all three classes of trans fatty acids raise the ratio of LDL to HDL. The effect of ruminant trans fatty acids and CLA on the LDL to HDL ratio was less than that of industrial trans fatty acids. The trans fatty acid with double bonds raised the LDL and lowered the HDL levels of cholesterol.         

            Thus, it was concluded that the removal of all the ruminants trans fatty acids (meat and milk) would lower the total trans fatty acid intake. Further studies need to be conducted to determine if the effects are due to chance. It some countries such as Denmark trans fatty acids are banned from the food industry.  

            This article helped me to better understand trans fatty acids to a larger extent. My knowledge of why trans fatty acids has such a negative impact on our bodies was broadened. Although this substance tantalized our taste buds and increases the shelf life of certain products it is a major component of cholesterol molecules. This as it is known leads to atherosclerosis which leads to heart attacks and strokes.

             I hope that after reading this  blog post you try to change your lifestyle to a more healthier way of living. Remember to exercise regularly, drink 6-8 glasses of water, include a large amount of fresh fruits and vegetables, and keep in mind that what you put into your body will affect you sooner or later.



Understanding Cholesterol…

What is Cholesterol?
Do you know what it is?
Have you ever stopped and wonder how many foods you eat each day that contains this molecule?

Well I am using this video to help you my viewers to understand more about this molecule.
Cholesterol is a waxy, fat like substance found in the blood stream and cells of the body. This naturally occurring substance plays a critical role in the formation of cell membranes and the manufacture of hormones. only a small amount of functions is needed to carry out these functions so the presence of additional cholesterol poses a risk to the body.

How cholesterol works?
This molecule does not dissolve in the blood stream but is transferred in and out of the cell by carriers called low density lipoproteins (LDL) and high density lipoproteins (HDL). When cholesterol increases more lipoproteins is needed to be produced to transfer it across the cell. LDLs are bad because too much of it result in plaque build up in the artery wall which leads to a condition know as atherosclerosis. The arteries are hardened and clogged which can lead to a heart attack or stroke. On the other hand HDL is a good carrier since it aids in the removal of cholesterol from the arteries into the liver and out of the body.

How is cholesterol determined in the body?
A blood test can be done. The levels of cholesterol varies in a person’s age, weight and sex.
An LDL level above 160 is high while an HDL level below 40 is too low. This places someone at risk for plaque build up. 75% of the cholesterol is made in the body while the other 25% is obtained from our diet. It is found in foods such as meat, eggs and liver. Eating less saturated fats from animals is a first step in lowering cholesterol levels and living a healthy life style.

This video was beneficial in helping me understand more about the effects cholesterol. A possible way to better this video is maybe the additional of more pictures to show the effects this molecule has on our body. I enjoyed it sice it was short and to the point.



Growing Up With Galactosemia

The video is about a young boy named Everet who was diagnosed with galactosemia at birth. He has grown up and is still able to survive with the help of his family and changing his lifestyle to accommodate this rare genetic disorder. He is unable to eat any dairy products such as milk, butter and cheese. He avoids foods high in galactose such as beans.
I have learnt more about this disease since it is the first time I ever heard about this disease and its effects.


Sugar is on my mind…haha
This video shows the ten steps in glycolyis. It deals with the breakdown of glucose into its various forms with the aid of the different enzymes to form pyruvate. Hope you enjoyed it like I did. :)


Lysine has three ionizable functional groups with the following pKa values:

α-amino group = 9.04

α-carboxylic group = 2.17

R group = 12.48

The R group for lysine is –CH2CH2CH2CH2NH2

(i) Write the equilibrium equations for its three ionizations and assign the proper pKa for each ionization. Show the net charge on the lysine molecule at each ionization stage.

lysine tit

(ii) Calculate the Isoelectric Point (pI):

The pI is the pH at which two amino acid has a net charge of zero (0).

1. Write out equations to show ionisation. Start with the amino acid in its fully protonated form. The pK values increase as you loose H-atoms so start the ionization using the group with the lowest pK value and then you increase as ionization continues.

2. Ensure that pk values are placed above reversible arrows.

3. Calculate the net charge of each molecule by adding the positive charges and subtracting the negative ones.

4. Calculate the pI using the equation below: ( pK1 value is the pk value to the left of the zero net charge and pK2 value is to the right of the net charge of zero).

pI = pk1 + pk2 ⁄ 2

pI = 9.04+12.48/2

pI = 10.76

All my Biochem Bloggers please note this titration steps for exam. It has been confirmed that it is going to come!!! Don’t say I didn’t warn