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Citric Acid: A key intermediate in metabolism. It is an acid compound found in citrus fruits. The salts of citric acid (citrates) can be used as anticoagulants due to their calcium chelating ability.

Products of the Citric Acid Cycle

JoVE 10977

The cells of most organisms—including plants and animals—obtain usable energy through aerobic respiration, the oxygen-requiring version of cellular respiration. Aerobic respiration consists of four major stages: glycolysis, pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation. The third major stage, the citric acid cycle, is also known as the Krebs cycle or tricarboxylic acid (TCA) cycle. For every glucose molecule that undergoes cellular respiration, the citric acid cycle is carried out twice; this is because glycolysis (the first stage of aerobic respiration) produces two pyruvate molecules per glucose molecule. During pyruvate oxidation (the second stage of aerobic respiration), each pyruvate molecule is converted into one molecule of acetyl-CoA—the input into the citric acid cycle. Therefore, for every glucose molecule, two acetyl-CoA molecules are produced. Each of the two acetyl-CoA molecules goes once through the citric acid cycle. The citric acid cycle begins with the fusion of acetyl-CoA and oxaloacetate to form citric acid. For each acetyl-CoA molecule, the products of the citric acid cycle are two carbon dioxide molecules, three NADH molecules, one FADH2 molecule, and one GTP/ATP molecule. Therefore, for every glucose molecule (which generates two acetyl-CoA molecules), the citric acid cycle yi

 Core: Biology

The Citric Acid Cycle

JoVE 10741

The citric acid cycle, also known as the Krebs cycle or TCA cycle, consists of several energy-generating reactions that yield one ATP molecule, three NADH molecules, one FADH2 molecule, and two CO2 molecules.

Acetyl CoA is the point-of-entry into the citric acid cycle, which occurs in the inner membrane (i.e., matrix) of mitochondria in eukaryotic cells or the cytoplasm of prokaryotic cells. Prior to the citric acid cycle, pyruvate oxidation produced two acetyl CoA molecules per glucose molecule. Hence, the citric acid cycle runs twice per glucose molecule. The citric acid cycle can be partitioned into eight steps, each yielding different molecules (italicized below). With the help of catalyzing enzymes, one acetyl CoA (2-carbon) reacts with oxaloacetic acid (4-carbon), forming the 6-carbon molecule citrate. Next, citrate is converted into one of its isomers, isocitrate, through a two-part process in which water is removed and added. The third step yields α-ketoglutarate (5-carbon) from oxidized isocitrate. This process releases CO2 and reduces NAD+ to NADH. The fourth step forms the unstable compound succinyl CoA from α-ketoglutarate, a process that also releases CO2 and reduces NAD+ to NADH. The fifth

 Core: Biology

Acid and Base Concentrations - Student Protocol

JoVE 11151

Preparation of ~0.1 M NaOH

In the first part of the lab, you will use a 50% w/w solution of NaOH to prepare 500 mL of ~0.1 M. The 50% w/w NaOH is indicative of its weight ratio. For example, if the instructor prepared 150 mL of the 50% w/w NaOH solution, then 150 g of NaOH was dissolved in 150 g of water, and the total weight of the solution is 300…

 Lab: Chemistry

Outcomes of Glycolysis

JoVE 11006

Nearly all the energy used by cells comes from the bonds that make up complex, organic compounds. These organic compounds are broken down into simpler molecules, such as glucose. Subsequently, cells extract energy from glucose over many chemical reactions—a process called cellular respiration.

Cellular respiration can take place in the presence or absence of oxygen, referred to as aerobic and anaerobic respiration, respectively. In the presence of oxygen, cellular respiration starts with glycolysis and continues with pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation. Both aerobic and anaerobic cellular respiration start with glycolysis. Glycolysis yields a net gain of two pyruvate molecules, two NADH molecules, and two ATP molecules (four produced minus two used during energy-requiring glycolysis). In addition to these major products, glycolysis generates two water molecules and two hydrogen ions. In cells that carry out anaerobic respiration, glycolysis is the primary source of ATP. These cells use fermentation to convert NADH from glycolysis back into NAD+, which is required to continue glycolysis. Glycolysis is also the primary source of ATP for mature mammalian red blood cells, which lack mitochondria. Cancer cells and stem cells rely on aerobic glycolysis for ATP. Cells that use aerobic respirati

 Core: Biology

Redox Reactions - Student Protocol

JoVE 11145

Making Solid Copper

In the first half of this lab, you will make solid copper. In this reaction, the solid copper is first oxidized by the strong oxidizing agent, nitric acid, to form water and the salt copper nitrate. The copper nitrate quickly dissociates into the ions, copper(2+) and nitrate, in water. The nitrate anions are reduced to NO2 gas,…

 Lab: Chemistry

Dietary Connections

JoVE 10746

Metabolic pathways are interconnected. The cellular respiration processes that convert glucose to ATP—such as glycolysis, pyruvate oxidation, and the citric acid cycle—tie into those that break down other organic compounds. As a result, various foods—from apples to cheese to guacamole—end up as ATP. In addition to carbohydrates, food also contains proteins and lipids—such as cholesterol and of these organic compounds are used as energy sources (i.e., to produce ATP). The human body possesses several enzymes that break down carbohydrates into simple sugars. While glucose can enter glycolysis directly, some simple sugars, such as fructose and galactose, are first converted into sugars that are intermediates of the glycolytic pathway. Proteins are broken down by enzymes into their constituent amino acids, which are usually recycled to create new proteins. However, if the body is starving or there is a surplus of amino acids, some amino acids can lose their amino groups and subsequently enter cellular respiration. The lost amino groups are converted into ammonia and incorporated into waste products. Different amino acids enter cellular respiration at different stages, including glycolysis, pyruvate oxidation, and the citric acid cycle. Amino acids can also be produced from intermediates in cellular respiration processes. Lipids, such as choleste

 Core: Biology

What is Cellular Respiration?

JoVE 10976

Organisms harvest energy from food, but this energy cannot be directly used by cells. Cells convert the energy stored in nutrients into a more usable form: adenosine triphosphate (ATP).

ATP stores energy in chemical bonds that can be quickly released when needed. Cells produce energy in the form of ATP through the process of cellular respiration. Although much of the energy from cellular respiration is released as heat, some of it is used to make ATP. During cellular respiration, several oxidation-reduction (redox) reactions transfer electrons from organic molecules to other molecules. Here, oxidation refers to electron loss and reduction to electron gain. The electron carriers NAD+ and FAD—and their reduced forms, NADH and FADH2, respectively—are essential for several steps of cellular respiration. Some prokaryotes use anaerobic respiration, which does not require oxygen. Most organisms use aerobic (oxygen-requiring) respiration, which produces much more ATP. Aerobic respiration generates ATP by breaking down glucose and oxygen into carbon dioxide and water. Both aerobic and anaerobic respiration begin with glycolysis, which does not require oxygen. Glycolysis breaks down glucose into pyruvate, yielding ATP. In the absence of oxygen, pyruvate ferments, producing NAD+ for continued glyc

 Core: Biology

Cellular Respiration- Concept

JoVE 10567

Autotrophs and Heterotrophs

Living organisms require a continuous input of energy to maintain cellular and organismal functions such as growth, repair, movement, defense, and reproduction. Cells can only use chemical energy to fuel their functions, therefore they need to harvest energy from chemical bonds of biomolecules, such as sugars and lipids. Autotrophic organisms, namely…

 Lab Bio

Electron Transport Chains

JoVE 10742

The final stage of cellular respiration is oxidative phosphorylation, which consists of (1) an electron transport chain and (2) chemiosmosis.

The electron transport chain is a set of proteins and other organic molecules found in the inner membrane of mitochondria in eukaryotic cells and the plasma membrane of prokaryotic cells. The electron transport chain has two primary functions: it produces a proton gradient—storing energy that can be used to create ATP during chemiosmosis—and generates electron carriers, such as NAD+ and FAD, that are used in glycolysis and the citric acid cycle. Generally, molecules of the electron transport chain are organized into four complexes (I-IV). The molecules pass electrons to one another through multiple redox reactions, moving electrons from higher to lower energy levels through the transport chain. These reactions release energy that the complexes use to pump H+ across the inner membrane (from the matrix into the intermembrane space). This forms a proton gradient across the inner membrane. NADH and FADH2 are reduced electron carriers produced during earlier cellular respiration phases. NADH can directly input electrons into complex I, which uses the released energy to pump protons into the intermembrane space. FADH2 inputs electrons into complex II, the only co

 Core: Biology


JoVE 10694

Mitochondria and peroxisomes are organelles that are the primary sites of oxygen usage in eukaryotic cells. Mitochondria carry out cellular respiration—the process that converts energy from food into ATP—the primary form of energy used by cells. Peroxisomes carry out a variety of functions, primarily breaking down different substances such as fatty acids.

Peroxisomes contain up to 50 enzymes and are surrounded by a single membrane. They carry out oxidative reactions that break down molecules and produce hydrogen peroxide (H2O2) as a by-product. H2O2 is toxic to cells, but the peroxisome contains an enzyme—catalase—that converts H2O2 into harmless water and oxygen. In addition, catalase uses H2O2 to break down alcohol in the liver into aldehyde and water. However, since H2O2 is produced in very low quantities in the body, other enzymes primarily degrade alcohol. A critical function of the peroxisome is to break down fatty acids in a process called β oxidation. The resulting product—acetyl-CoA—is released into the cytosol and can travel to the mitochondria, where it is used to produce ATP. In mammalian cells, the mitochondria also carry out β oxidation, as well as using products from the catabolism o

 Core: Biology

ATP Yield

JoVE 11008

Cellular respiration produces 30-32 ATP molecules per glucose molecule. Although most of the ATP results from oxidative phosphorylation and the electron transport chain (ETC), 4 ATP are gained beforehand (2 from glycolysis and 2 from the citric acid cycle).

The ETC is embedded in the inner mitochondrial membrane and comprises four main protein complexes and an ATP synthase. NADH and FADH2 pass electrons to these complexes, which in turn pump protons into the intermembrane space. This distribution of protons generates a concentration gradient across the membrane. The gradient drives the production of ATP when protons flow back into the mitochondrial matrix via the ATP synthase. For every 2 input electrons that NADH passes into complex I, complexes I and III each pump 4 protons and complex IV pumps 2 protons, totaling 10 protons. Complex II is not involved in the electron chain initiated by NADH. FADH2, however, passes 2 electrons to complex II, so a total of 6 protons are pumped per FADH2; 4 protons via complex III and 2 via complex IV. Four protons are needed to synthesize 1 ATP. Since 10 protons are pumped for every NADH, 1 NADH yields 2.5 (10/4) ATP. Six protons are pumped for every FADH2, so 1 FADH2 yields 1.5 (6/4) ATP. Cellular respiration produces a maximum of 10 NADH and 2 FADH2

 Core: Biology

Pyruvate Oxidation

JoVE 10740

After glycolysis, the charged pyruvate molecules enter the mitochondria via active transport and undergo three enzymatic reactions. These reactions ensure that pyruvate can enter the next metabolic pathway so that energy stored in the pyruvate molecules can be harnessed by the cells.

First, the enzyme pyruvate dehydrogenase removes the carboxyl group from pyruvate and releases it as carbon dioxide. The stripped molecule is then oxidized and releases electrons, which are then picked up by NAD+ to produce NADH, forming acetate. Finally, coenzyme A—a sulfur-containing compound derived from a B vitamin—attaches to the acetate via its sulfur atom to create acetyl coenzyme A, or acetyl CoA. Acetyl CoA then moves into the citric acid cycle where it will be further oxidized.

 Core: Biology

What is Glycolysis?

JoVE 10737

Cells make energy by breaking down macromolecules. Cellular respiration is the biochemical process that converts “food energy” (from the chemical bonds of macromolecules) into chemical energy in the form of adenosine triphosphate (ATP). The first step of this tightly regulated and intricate process is glycolysis. The word glycolysis originates from Latin glyco (sugar) and lysis (breakdown). Glycolysis serves two main intracellular functions: generate ATP and intermediate metabolites to feed into other pathways. The glycolytic pathway converts one hexose (six-carbon carbohydrate such as glucose), into two triose molecules (three-carbon carbohydrate) such as pyruvate, and a net of two molecules of ATP (four produced, two consumed) and two molecules of nicotinamide adenine dinucleotide (NADH). Did you know that glycolysis was the first biochemical pathway discovered? In the mid-1800s, Louis Pasteur determined that microorganisms cause the breakdown of glucose in the absence of oxygen (fermentation). In 1897, Eduard Buchner found that fermentation reactions can still be carried out in cell-free yeast extracts, achieved by breaking open the cell and collecting the cytoplasm which contains the soluble molecules and organelles. Shortly thereafter in 1905, Arthur Harden and William Young discovered that the rate of fermentation decreases wit

 Core: Biology

Electron Carriers

JoVE 10744

Electron carriers can be thought of as electron shuttles. These compounds can easily accept electrons (i.e., be reduced) or lose them (i.e., be oxidized). They, therefore, play an essential role in energy production because cellular respiration is contingent on the flow of electrons.

Over the many stages of cellular respiration, glucose breaks down into carbon dioxide and water. Electron carriers pick up electrons lost by glucose in these reactions, temporarily store the electrons and input them into the electron transport chain. Two such electron carriers are NAD+ and FAD, which are both derived from B vitamins. The reduced forms of NAD+ and FAD, NADH and FADH2, respectively, are produced during earlier stages of cellular respiration (glycolysis, pyruvate oxidation, and the citric acid cycle). The reduced electron carriers NADH and FADH2 pass electrons into complexes I and II of the electron transport chain, respectively. In the process, they are oxidized to form NAD+ and FAD. Additional electron carriers in the electron transport chain are flavoproteins, iron-sulfur clusters, quinones, and cytochromes. With the assistance of enzymes, these electron carriers eventually transfer the electrons to oxygen molecules. The electron carriers become oxidized as they donate electrons and re

 Core: Biology

Redox Reactions - Prep Student

JoVE 11143

Preparation of Solutions

Here, we show the laboratory preparation for 10 students working in pairs, with some excess. Please adjust quantities as needed.

To set up for this lab experiment, wear the appropriate personal protective equipment, including a lab coat, chemical splash goggles, and gloves. All solutions …

 Lab: Chemistry

High-Performance Liquid Chromatography (HPLC)

JoVE 10156

Source: Dr. Paul Bower - Purdue University

High-performance liquid chromatography (HPLC) is an important analytical method commonly used to separate and quantify components of liquid samples. In this technique, a solution (first phase) is pumped through a column that contains a packing of small porous particles with a second phase bound to …

 Analytical Chemistry

Efficiency of Liquid-liquid Extraction

JoVE 10426

Source: Kerry M. Dooley and Michael G. Benton, Department of Chemical Engineering, Louisiana State University, Baton Rouge, LA

Liquid-liquid extraction (LLE) is a separation technique used instead of distillation when either: (a) the relative volatilities of the compounds to be separated are very similar; (b) one or more of the mixture…

 Chemical Engineering


JoVE 10743

Oxidative phosphorylation is a highly efficient process that generates large amounts of adenosine triphosphate (ATP), the basic unit of energy that drives many processes in living cells. Oxidative phosphorylation involves two processes—electron transport and chemiosmosis. During electron transport, electrons are shuttled between large complexes on the inner mitochondrial membrane and protons (H+) are pumped across the membrane into the intermembrane space, creating an electrochemical gradient. In the next step, protons flow back down their gradient into the mitochondrial matrix via ATP synthase, a protein complex embedded within the inner membrane. This process, called chemiosmosis, uses the energy of the proton gradient to drive the synthesis of ATP from adenosine diphosphate (ADP). The electron transport chain is a series of complexes that transfer electrons from electron donors to electron acceptors via simultaneous reduction and oxidation reactions, otherwise known as redox reactions. At the end of the chain, electrons reduce molecular oxygen to produce water. The shuttling of electrons between complexes is coupled with proton transfer, whereby protons (H+ ions) travel from the mitochondrial matrix to the intermembrane space against their concentration gradient. Eventually, the high concentration of protons in the interm

 Core: Biology

VirWaTest, A Point-of-Use Method for the Detection of Viruses in Water Samples

1Laboratory of Viruses Contaminants of Water and Food (VIRCONT), Department of Genetics, Microbiology and Statistics, Section of Microbiology, Virology and Biotechnology, University of Barcelona, 2Grupo de Investigación Biodiversidad, Medio Ambiente y Salud (BIOMAS), Facultad de Ingenierías y Ciencias Agropecuarias (FICA), Ingeniería en Biotecnología, Universidad de las Américas, 3Municipal Laboratory - Waters of Mataró, 4GenIUL

JoVE 59463

 Immunology and Infection

Construction of Models for Nondestructive Prediction of Ingredient Contents in Blueberries by Near-infrared Spectroscopy Based on HPLC Measurements

1United Graduate School of Agricultural Science, Tokyo University of Agriculture and Technology, 2Faculty of Agriculture, Tokyo University of Agriculture and Technology, 3Institute of Agriculture, Tokyo University of Agriculture and Technology

JoVE 53981


Phage Phenomics: Physiological Approaches to Characterize Novel Viral Proteins

1Department of Biology, San Diego State University, 2Computational Science Research Center, San Diego State University, 3Bioinformatics and Medical Informatics Research Center, San Diego State University, 4Department of Mathematics and Statistics, San Diego State University, 5Department of Computer Science, San Diego State University, 6Mathematics and Computer Science Division, Argonne National Laboratory, 7SPARC Committee, Broad Institute

JoVE 52854

 Immunology and Infection

High Efficiency Differentiation of Human Pluripotent Stem Cells to Cardiomyocytes and Characterization by Flow Cytometry

1Department of Biochemistry, Medical College of Wisconsin, 2Stanford Cardiovascular Institute, Stanford University School of Medicine, 3Department of Anesthesiology, Medical College of Wisconsin, 4Stem Cell and Regenerative Medicine Consortium, LKS Faculty of Medicine, Hong Kong University, 5Division of Cardiology, Johns Hopkins University School of Medicine, 6Cardiovascular Research Center, Biotechnology and Bioengineering Center, Medical College of Wisconsin

JoVE 52010


Pancreatic Tissue-Derived Extracellular Matrix Bioink for Printing 3D Cell-Laden Pancreatic Tissue Constructs

1Department of Mechanical Engineering, Pohang University of Science and Technology, 2School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, 3Asan Institute for Life Science, University of Ulsan College of Medicine and Asan Medical Center, 4Division of Hepato-Biliary and Pancreatic Surgery, Department of Surgery, University of Ulsan College of Medicine and Asan Medical Center, 5Department of Creative IT Engineering, Pohang University of Science and Technology

JoVE 60434


Genetic Manipulation of the Plant Pathogen Ustilago maydis to Study Fungal Biology and Plant Microbe Interactions

1Institute for Microbiology, Heinrich-Heine University Düsseldorf, 2Bioeconomy Science Center (BioSC), 3Department of Genetics, Institute of Applied Biosciences, Karlsruhe Institute of Technology, 4Cluster of Excellence in Plant Sciences (CEPLAS), Heinrich-Heine University Düsseldorf

JoVE 54522


High Throughput Traction Force Microscopy Using PDMS Reveals Dose-Dependent Effects of Transforming Growth Factor-β on the Epithelial-to-Mesenchymal Transition

1Department of Bioengineering, McGill University, 2Goodman Cancer Research Centre, McGill University, 3Department of Medicine, McGill University, 4Department of Emergency Medicine, Beth Israel Deaconess Medical Center

JoVE 59364

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