If you have been reading about Freeze-thaw and want a single page that covers the useful parts, this is it: definitions, context, how it is studied, and the questions that come up repeatedly.
Updated 2026-04-22. Numbers and descriptions here follow the published literature rather than marketing material.
Beyond redox chemistry, NAD+ serves as a substrate for enzymes that cleave the molecule and transfer its ADP-ribose moiety or remove acetyl groups. Sirtuins consume NAD+ during deacetylation, poly(ADP-ribose) polymerases use it in DNA damage responses, and CD38 enzymes hydrolyze it to signaling metabolites. These consumption pathways mean that NAD+ availability can influence gene regulation, DNA repair, and calcium signaling. Cellular NAD+ concentrations decline in some tissues with age in animal models, but whether this decline is a cause or consequence of aging in humans remains an active open question.
Nicotinamide adenine dinucleotide, commonly abbreviated NAD+, is a dinucleotide coenzyme built from an adenine nucleotide and a nicotinamide nucleotide joined by a pyrophosphate linkage. Its oxidized form carries a positive charge on the nicotinamide ring, while the reduced form, NADH, carries a hydride equivalent. The molecule participates in hundreds of oxidoreductase reactions, where it accepts or donates electrons and protons. Because it can cycle between oxidized and reduced states without net consumption, NAD+ functions as a reusable electron carrier rather than a fuel molecule.
In glycolysis, the tricarboxylic acid cycle, and fatty acid oxidation, NAD+ is reduced to NADH at specific dehydrogenase steps. NADH then delivers electrons to the mitochondrial electron transport chain, mainly at complex I, supporting oxidative phosphorylation and ATP production. The balance between NAD+ and NADH, often expressed as a ratio, influences metabolic flux and redox homeostasis in different cellular compartments. Cytosolic and mitochondrial pools are connected but not identical, and their ratios can differ substantially because of compartment-specific enzymes and transport systems.
Commercial NAD+ is supplied as a solid, often as the free acid or a salt, and purity is verified by chromatographic methods. Laboratories typically store it desiccated at minus 20 degrees Celsius or below. Working solutions are prepared fresh because even sterile aqueous solutions can lose activity over hours to days depending on pH and temperature. Documentation may include a certificate of analysis, an assay value, and a recommended retest date. Researchers should verify identity and purity when results depend on precise cofactor concentrations.
NAD+ is commonly measured by high-performance liquid chromatography with ultraviolet detection, often at 254 or 260 nm. Enzymatic cycling assays provide higher sensitivity by coupling NAD+ to a reporter reaction. Mass spectrometry can distinguish NAD+ from close analogues and confirm isotope labeling. Sample preparation usually involves rapid quenching of metabolism to prevent interconversion with NADH. Because NAD+ and NADH differ by one hydride, extraction conditions strongly affect the measured ratio.
In aqueous solution, NAD+ is most stable under mildly acidic to neutral conditions and degrades faster at high pH or elevated temperature. The molecule can hydrolyze at the pyrophosphate bond or undergo nonenzymatic cyclization. Buffers, chelating agents, and cold temperatures slow these losses during analysis. Repeated freeze-thaw cycles are generally avoided because they can promote degradation and concentration changes. Light exposure is also controlled, though NAD+ is less photolabile than some related nucleotides.
| Property | Value | Notes |
|---|---|---|
| Common synonyms | β-NAD+, coenzyme I, DPN | DPN stands for diphosphopyridine nucleotide; older literature uses this term. |
| CAS Registry Number | 53-84-9 | Free acid form of β-nicotinamide adenine dinucleotide. |
| Molecular formula | C21H27N7O14P2 | Anhydrous free acid; molar mass 663.43 g/mol. |
| Appearance | White to off-white powder | Crystalline solid; may absorb moisture from air. |
| Solubility | Freely soluble in water | Insoluble in most nonpolar organic solvents. |
Beyond redox chemistry, NAD+ is consumed as a substrate by enzymes that transfer ADP-ribose or remove acetyl groups. Sirtuins use NAD+ in deacylation reactions, poly(ADP-ribose) polymerases use it in DNA damage responses, and CD38 hydrolases convert it to signaling metabolites. Because these enzymes compete for the same pool, changes in NAD+ availability can influence multiple cellular processes. The relative contribution of each consumption route differs by cell type and condition, and precise quantitative links remain an active area of study.
Research on NAD+ spans biochemistry, aging biology, and metabolism. Studies often examine how NAD+ levels change with age, diet, exercise, or disease states, and whether precursor supplementation alters those levels. Findings in animal models do not automatically translate to humans, and measurement methods vary across studies. Questions about tissue-specific effects, long-term consequences, and causal relationships remain open. NAD+ itself is not established as a single therapeutic agent with a broad clinical role.
Nicotinamide adenine dinucleotide, abbreviated NAD+, is a dinucleotide composed of two nucleotides joined by phosphate groups. One nucleotide contains adenine; the other contains nicotinamide. The molecule exists in oxidized (NAD+) and reduced (NADH) forms, and the reversible hydride transfer between them underlies many metabolic oxidation-reduction reactions. In cells, NAD+ serves as an electron acceptor in pathways such as glycolysis, the citric acid cycle, and oxidative phosphorylation. Its concentration and redox ratio vary by compartment, tissue, and metabolic state.
Biosynthesis of NAD+ starts from nicotinamide, nicotinic acid, or nicotinamide riboside through salvage pathways. A rate-limiting enzyme, nicotinamide phosphoribosyltransferase, converts nicotinamide to nicotinamide mononucleotide. Further coupling with ATP yields NAD+. In mammals, the liver and muscle can synthesize NAD+ from dietary precursors, but tissue levels vary widely. Researchers study these pathways to understand age-related changes, metabolic disorders, and neurodegeneration. Direct causal links between NAD+ decline and disease remain an active area of investigation.
NAD+ is a dinucleotide composed of adenine, ribose, and nicotinamide groups joined by phosphate linkages. It serves as a coenzyme in oxidoreductase reactions, cycling between oxidized NAD+ and reduced NADH. The molecule is water-soluble and occurs in all living cells. Its nicotinamide ring accepts hydride ions during catabolic reactions, linking substrate oxidation to electron transport. This redox couple supports ATP production and helps maintain cytosolic and mitochondrial redox balance in many cell types.
The molecule was first described in the early twentieth century as a factor that promoted fermentation in yeast extracts. Later work linked it to hydrogen transfer and to the oxidation of nutrients in living tissues. Its structure was resolved as a dinucleotide, which explained why it could accept and donate electrons at specific enzyme sites. Today, NAD+ is recognized as a central substrate and signaling precursor, not merely a metabolic cofactor. Whether all observed NAD+ changes reflect causal signaling remains an open question.
Related compounds include NADH, the reduced form, and NADP+, which carries an additional phosphate group. NADP+ and NADPH often serve in biosynthetic and antioxidant reactions, while NAD+ and NADH are more associated with energy-yielding catabolism. Nicotinamide, nicotinic acid, and nicotinamide riboside are precursors that can enter salvage pathways. The exact contribution of dietary precursors to tissue NAD+ pools is an area of active investigation. Some studies measure labeled precursors to trace those routes.
Beyond electron transfer, NAD+ serves as a substrate for enzymes that cleave the molecule and transfer ADP-ribose units. Sirtuins, poly(ADP-ribose) polymerases, and CD38 consume NAD+ in regulatory reactions. These activities link NAD+ availability to DNA repair, chromatin modification, calcium signaling, and metabolic stress responses. Because consumption can exceed biosynthesis under some conditions, cellular NAD+ levels are dynamic rather than fixed. Enzyme affinity and local synthesis also influence how much NAD+ is available for signaling.
NAD+ is a dinucleotide composed of adenine, ribose, and nicotinamide moieties linked by phosphate groups. Its oxidized form carries a positive charge on the nicotinamide ring, which enables reversible hydride transfer. The molecule functions as a coenzyme in oxidoreductase reactions rather than as a dietary vitamin in its intact form. Cells maintain separate pools in cytoplasm, mitochondria, and nucleus. This compartmentalization allows distinct redox environments while preserving a shared chemical identity.
The first steps to digitization in trucking came in the form of digital freight exchanges such as Teleroute and TIMOCOM. Greater efficiency in trucking is being achieved through intelligent freight technology such as automated interfaces that can help truckers and regulatory bodies reduce stops at weigh stations and time spent at border checkpoints. This improves schedule adherence, reduces administrative burdens, and increases fuel efficiency by minimizing time spent idling. Mobile tracking programs can also reduce theft and loss of cargo by recording instances where trailer doors are opened outside of approved areas (geo-fences) and alerting authorities. Growing automation of quoting and booking of truckload shipments and increased automation of matching trucks with appropriate loads in a timely manner is also working to maximize efficiency for shippers and consumers. Cargofy is an example of a freight technology company using artificial intelligence to automate freight procurement, load matching, dispatching and other logistics operations. Vehicle tracking has been used by shipping agents and freight forwarders to monitor the location of vehicles. They can also monitor traffic information, vehicle and driver data, and other real-time freight information. Due to the relatively short life cycle of three to four years for commercial trucks, implementation of new interfaces and freight technologies has progressed more quickly in trucking than in other sectors.
The Straw Hats' Jolly Roger (Japanese: 麦わらの一味の海賊旗, Hepburn: Mugiwara no Ichimi no Kaizokuki; lit. Straw Hat Crew's Jolly Roger), also known as the Straw Hats' Skull and Crossbones, is a pirate flag predominantly featured in the Japanese One Piece manga and its anime franchise, wherein it is used to identify the protagonist Straw Hats, a pirate crew who often fight for liberation of oppressed peoples. The fictional Jolly Roger is of a cartoonish skull with two crossbones and a straw hat similar to the one worn by Monkey D. Luffy. The Straw Hats' Jolly Roger is a symbol for several global youth-led protest movements. In the August 2025 Indonesian protests, people began using the flag instead of the national flag as a form of protest against the government, and the practice spread to other countries. Notably, the flag saw widespread use during protests in Nepal and Madagascar which overthrew their respective governments. The flag has also seen use in protests in other countries across Africa, the Americas, and Eurasia.
== Treatment == During the early 1940s, rabbit models in combination with the drug penicillin allowed for a long-term drug treatment. These experiments established the groundwork that modern scientists use for syphilis therapy. Penicillin can inhibit T. pallidum in 6–8 hours, though the cells still remain in lymph nodes and regenerate. Penicillin is not the only drug that can be used to inhibit T. pallidum; any β-lactam antibiotics or macrolides can be used. The T. pallidum strain 14 has built-in resistance to some macrolides, including erythromycin and azithromycin. Resistance to macrolides in T. pallidum strain 14 is believed to derive from a single-point mutation that increased the organism's livability. Many of the syphilis treatment therapies only lead to bacteriostatic results, unless larger concentrations of penicillin are used for bactericidal effects. Penicillin overall is the most recommended antibiotic by the Centers for Disease Control, as it shows the best results with prolonged use. It can inhibit and may even kill T. pallidum at low to high doses, with each increase in concentration being more effective. The Guideline Development Group has recommended the development of a new treatment, a short course treatment that is administered orally and can cross the placental barriers in pregnant women.
== Further reading == Arthur, Joyce (1996). "Creationism: Bad Science or Immoral Pseudoscience?". Skeptic. 4 (4): 88–93. Archived from the original on March 7, 2012. Montagu, M. F. Ashley, ed. (1984-01-12). Science and Creationism (Kindle ed.). USA: Oxford University Press. ASIN B000QXDJIG. Nelkin, Dorothy (2000) [1986]. The Creation Controversy: Science or Scripture in the Schools. toExcel. ISBN 0-595-00194-7. Numbers, Ronald L. (2006). The Creationists: From Scientific Creationism to Intelligent Design (expanded ed.). Harvard. ISBN 978-0-674-02339-0.
=== TNXB (functional gene) === The TNXB gene localizes to the major histocompatibility complex (MHC class III) region on chromosome 6. The structure of this gene is unusual in that it overlaps the CREBL1 and CYP21A2 genes at its 5' and 3' ends, respectively.
Sources: en.wikipedia.org
Pyridinoline, also known as Hydroxylysylpyridinoline, is a fluorescent cross-linking compound of collagen fibers. Crosslinks in collagen and elastin are derived from lysyl and hydroxylysyl residues, a process catalyzed by lysyl oxidase. Fujimoto and colleagues first described the isolation and characterization of a fluorescent material in bovine Achilles tendon collagen and termed it pyridinoline. It is reported to be present in collagen of bone and cartilage, but is absent in collagen of skin. It is not present in newly synthesized collagen and is formed from aldimine cross-links during maturation of collagen fibers. Pyridinoline and deoxypyridinoline were found to be released into the blood during bone degradation and rapidly excreted in the urine. In a preliminary study, both these compounds were proposed as a marker for metastatic bone tumor in patients with prostate cancer.
Lutetium (177Lu), which decays with the emission of a low-energy electron, is useful for treating some cancerous tumors. Several compounds of 177Lu are available for this purpose: 1) Lutetium chloride, LuCl3, sold under the brand name Lumark among others, is used for radiolabeling other medicines, either as an anti-cancer therapy or for scintigraphy (medical radio-imaging). Its most common side effects are anaemia (low red blood cell counts), thrombocytopenia (low blood platelet counts), leucopenia (low white blood cell counts), lymphopenia (low levels of lymphocytes, a particular type of white blood cell), nausea (feeling sick), vomiting and mild and temporary hair loss. 2) Lutetium (177Lu) oxodotreotide, also known as Lutetium (177Lu) dotatate ("Lutathera"), has been approved for some types of tumors. 3) Lutetium (177Lu) vipivotide tetraxetan ("Pluvicto") has also been approved.
=== Perception === Machine perception is the ability to use input from sensors (such as cameras, microphones, wireless signals, active lidar, sonar, radar, and tactile sensors) to deduce aspects of the world. Computer vision is the ability to analyse visual input. The field includes speech recognition, image classification, facial recognition, object recognition, object tracking, and robotic perception.
Beneš—the leader of the Czechoslovak government-in-exile—and František Moravec—head of Czechoslovak military intelligence—organized and coordinated a resistance network. Hácha, Prime Minister Alois Eliáš, and the Czechoslovak resistance acknowledged Beneš's leadership. Active collaboration between London and the Czechoslovak home front was maintained throughout the war years. The most important event of the resistance was Operation Anthropoid, the assassination of Reinhard Heydrich, SS leader Heinrich Himmler's deputy and the then Protector of Bohemia and Moravia. Infuriated, Hitler ordered the arrest and execution of 10,000 randomly selected Czechs. Over 10,000 were arrested, and at least 1,300 were executed. According to one estimate, 5,000 were killed in reprisals. The assassination resulted in one of the most well-known reprisals of the war. The Nazis completely destroyed the villages of Lidice and Ležáky; all men over 16 years from the village were murdered, and the rest of the population was sent to Nazi concentration camps where many women and nearly all the children were killed. The Czechoslovak resistance comprised four main groups:
Sources: en.wikipedia.org
Linus Carl Pauling was born on February 28, 1901, in Portland, Oregon, the firstborn child of Herman Henry William Pauling (1876–1910) and Lucy Isabelle "Belle" Darling (1881–1926). He was named "Linus Carl", in honor of Lucy's father, Linus, and Herman's father, Carl. His ancestry included German and English-Scottish. In 1902, after his sister Pauline was born, Pauling's parents decided to move out of Portland to find more affordable and spacious living quarters than their one-room apartment. Lucy stayed with her husband's parents in Lake Oswego until Herman brought the family to Salem, where he worked briefly as a traveling salesman for the Skidmore Drug Company. Within a year of Lucile's birth in 1904, Herman Pauling moved his family to Lake Oswego, Oregon where he opened his own drugstore. He moved his family to Condon, Oregon, in 1905. By 1906, Herman Pauling was suffering from recurrent abdominal pain. He died of a perforated ulcer on June 11, 1910, leaving Lucy to care for Linus, Lucile and Pauline. Pauling attributes his interest in becoming a chemist to being amazed by experiments conducted by a friend, Lloyd A. Jeffress, who had a small chemistry lab kit. He later wrote: "I was simply entranced by chemical phenomena, by the reactions in which substances, often with strikingly different properties, appear; and I hoped to learn more and more about this aspect of the world." In high school, Pauling conducted chemistry experiments by scavenging equipment and material from an abandoned steel plant.
==== Israel ==== Dunkin' Donuts Israel (Hebrew: דאנקן דונאטס) was an Israeli franchise of the chain. Dunkin' Donuts Israel opened their first location in Israel in 1996 in Tel Aviv. Their main flagship store was located in Rabin Square, with their factory in Lod. Upon opening their first location, Dunkin' was a huge hit. When the original location opened, it broke Dunkin's own sales records by selling 3 million doughnuts in the first eight months. The chain's manager was quoted saying "Israelis do not stop eating doughnuts, They buy such quantities, quantities that we would never have believed they would buy. They buy boxes of 12, eat them here, and then buy more boxes to take home." In 2001, when Dunkin' Donuts Israel decided to close, the company had accumulated a large amount of debt. When the Second Intifada broke out, Israeli tourism slowed, and Dunkin' decided to close. Approximately $2 million was invested in Dunkin' Donuts Israel when it failed. At the peak, Dunkin' Donuts had 9 branches in Israel, short of their original goal of 15, including 3 in Jerusalem.
Citizens may teach and may be taught religion privately.' Among further restrictions, those adopted in 1929 included express prohibitions on a range of church activities, including meetings for organized Bible study. Both Christian and non-Christian establishments were shut down by the thousands in the 1920s and 1930s. By 1940, as many as 90% of the churches, synagogues, and mosques that had been operating in 1917 were closed; the majority of them were demolished or re-purposed for state needs with little concern for their historic and cultural value. More than 85,000 Orthodox priests were shot in 1937 alone. Only a twelfth of the Russian Orthodox Church's priests were left functioning in their parishes by 1941. In the period between 1927 and 1940, the number of Orthodox Churches in Russia fell from 29,584 to less than 500 (1.7%). The Soviet Union was officially a secular state, but a 'government-sponsored program of forced conversion to atheism' was conducted under the doctrine of state atheism. The government targeted religions based on state interests, and while most organized religions were never outlawed, religious property was confiscated, believers were harassed, and religion was ridiculed while atheism was propagated in schools. In 1925, the government founded the League of Militant Atheists to intensify the propaganda campaign.
In patients with the retinal disorder, neovascular age-related macular degeneration, local anti-VEGF therapy to limit the bio-activity of vascular endothelial growth factor has been shown to protect vision by limiting progression. In a wide range of cancers, treatment approaches have been studied, or are in development, aimed at decreasing tumour growth by reducing angiogenesis.
The problem can be overcome by stopping the reaction with sodium hydroxide (NaOH) or other strong base, which converts all product into 4-nitrophenoxide; final pH must be > ca. 9.2 to ensure more than 99% of the product is ionised. Alternatively enzyme activity can be measured at 348 nm, the isosbestic point for 4-nitrophenol/4-nitrophenoxide.
Sources: en.wikipedia.org
NAD+ is an oxidized dinucleotide coenzyme that carries electrons in metabolic reactions. It is also consumed by signaling enzymes, including sirtuins and PARPs. Its reduced form is NADH.
NAD+ is the oxidized form and can accept a hydride equivalent. NADH is the reduced form and donates electrons to the electron transport chain. The two forms cycle between each other during cellular respiration.
In mammals, NAD+ is synthesized mainly through salvage pathways using nicotinamide, nicotinamide riboside, or nicotinic acid. Tryptophan can also contribute through a de novo route. The salvage pathway is often considered the primary source in many tissues.
NAD+ and NADH can interconvert quickly after a sample is collected, which can alter the measured ratio. Rapid quenching and cold handling limit enzymatic and chemical changes.