l-ascorbic acid

Electron Donation and Redox Potential in Enzyme Catalysis

This compound is a potent reducing agent, readily donating electrons to other molecules. oregonstate.edu This capacity is fundamental to its function in enzymatic reactions. The molecule can donate two electrons sequentially. nih.gov The loss of the first electron results in the formation of the relatively stable ascorbyl radical (semidehydroascorbic acid). nih.govresearchgate.net This ability to donate electrons allows this compound to reduce reactive oxygen species and other harmful free radicals, thereby protecting cellular components from oxidative damage. researchgate.net In the context of enzyme catalysis, this electron-donating property is harnessed to regenerate the active form of metalloenzymes. oregonstate.edubrieflands.com

The redox potential of this compound makes it an effective reducing agent in numerous enzymatic reactions. researchgate.net It can reduce metal ions, such as iron and copper, which are essential components of many enzymes. researchgate.netencyclopedia.pub This reduction is crucial for the catalytic cycle of these enzymes to proceed. lumenlearning.com

Role in Maintaining Enzyme-Bound Metals in Reduced Forms

Many enzymes, particularly mixed-function oxidases, rely on metal ions like iron (Fe) and copper (Cu) at their active sites. oregonstate.eduresearchgate.net These enzymes catalyze hydroxylation reactions that are vital for the synthesis of various biomolecules. oregonstate.edu During the catalytic cycle, the metal ion can become oxidized, rendering the enzyme inactive. lumenlearning.com For instance, in the case of iron-containing dioxygenases, the ferrous iron (Fe²⁺) at the active site can be oxidized to ferric iron (Fe³⁺). lumenlearning.com

This compound acts as a specific reductant to regenerate the active form of these enzymes by reducing the oxidized metal ion back to its active, reduced state. oregonstate.edubrieflands.comlumenlearning.com For example, it reduces Fe³⁺ back to Fe²⁺, allowing the enzyme to participate in another round of catalysis. lumenlearning.com This function is essential for the continuous activity of enzymes like prolyl and lysyl hydroxylases, which are critical for collagen synthesis. lumenlearning.comnih.govnews-medical.net Without this compound, these enzymes would be rapidly inactivated, leading to impaired collagen production. lumenlearning.com

Lysyl Hydroxylases

Criticality in Collagen Cross-link Formation

This compound is indispensable for the biosynthesis of collagen, the most abundant protein in mammals. jddtonline.info Its role is particularly critical for the function of lysyl hydroxylase, an enzyme essential for the formation of stable collagen cross-links. nih.gov Lysyl hydroxylase catalyzes the hydroxylation of lysine residues within procollagen chains to form hydroxylysine. lumenlearning.com This hydroxylation is a prerequisite for the subsequent glycosylation and the formation of covalent cross-links between collagen molecules. lumenlearning.com These cross-links are vital for the tensile strength and stability of collagen fibers, which provide structural integrity to connective tissues throughout the body. lumenlearning.com

In the absence of sufficient this compound, the activity of lysyl hydroxylase is impaired, leading to under-hydroxylation of lysine residues. lumenlearning.com This results in the production of weak, unstable collagen that is unable to form proper cross-links and is more susceptible to degradation. lumenlearning.com The consequence is a weakening of connective tissues, a hallmark of scurvy.

Stimulation of Lysyl Hydroxylase Activity

Beyond its role as a cofactor, this compound has been shown to directly stimulate the activity of lysyl hydroxylase. nih.gov Studies on cultured human skin fibroblasts have demonstrated that the addition of this compound to the culture medium leads to an increase in lysyl hydroxylase activity. lorealdermatologicalbeauty.com This stimulatory effect appears to be independent of its role in maintaining the iron cofactor in a reduced state. The increased activity of lysyl hydroxylase in the presence of this compound contributes to enhanced collagen production and the formation of a stable connective tissue matrix. researchgate.net Interestingly, research has also indicated that this compound can inhibit the activity of prolyl hydroxylase in the same cell type, suggesting a differential regulation of these two important collagen-modifying enzymes. nih.govlorealdermatologicalbeauty.com Some studies have also suggested that this compound may inhibit the activity of lysyl oxidase, another enzyme involved in a later stage of collagen cross-linking. tandfonline.comnih.gov

Carnitine Biosynthesis Enzymes

This compound serves as a vital cofactor for two key enzymes in the carnitine biosynthesis pathway: ε-N-trimethyllysine hydroxylase and γ-butyrobetaine hydroxylase. nih.govmdpi.com Carnitine is essential for the transport of long-chain fatty acids into the mitochondria for beta-oxidation, a critical process for energy production. mdpi.comliposhell.pl Both ε-N-trimethyllysine hydroxylase and γ-butyrobetaine hydroxylase are α-ketoglutarate-dependent dioxygenases that require iron and this compound to function optimally. nih.gov this compound maintains the iron in the active sites of these enzymes in its reduced ferrous (Fe²⁺) state. nutri-facts.org A deficiency in this compound can impair the activity of these enzymes, leading to a decrease in the rate of carnitine synthesis. nih.gov

Neuropeptide Amidation (Peptidylglycine alpha-amidating monooxygenase)

Many neuropeptides and peptide hormones require a C-terminal amide group for their full biological activity. nih.govnih.gov This amidation is catalyzed by the bifunctional enzyme peptidylglycine alpha-amidating monooxygenase (PAM). nih.gov The first step of this two-step reaction is catalyzed by the peptidyl-alpha-hydroxylating monooxygenase (PHM) domain of PAM, which is a copper- and this compound-dependent enzyme. nih.govuniprot.org PHM hydroxylates the alpha-carbon of the C-terminal glycine residue of a peptide substrate. uniprot.org this compound is essential for this reaction, acting as an electron donor to maintain the copper ions in the enzyme's active site in a reduced state. nih.gov The subsequent step is catalyzed by the peptidyl-α-hydroxyglycine α-amidating lyase (PAL) domain, which cleaves the modified glycine to produce the amidated peptide and glyoxylate. uniprot.org The availability of this compound can therefore be a rate-limiting factor in the biosynthesis of numerous active neuropeptides and hormones. nih.gov

Catecholamine Biosynthesis (Dopamine β-hydroxylase)

This compound is a critical cofactor for dopamine β-hydroxylase (DBH), a copper-containing monooxygenase enzyme that catalyzes the conversion of dopamine to norepinephrine. wikipedia.orgmdpi.comwikipedia.org This reaction is a crucial step in the biosynthesis of catecholamines, a group of neurotransmitters that includes dopamine, norepinephrine, and epinephrine. mdpi.com The synthesis of these neurotransmitters is vital for proper brain function and mood regulation. patsnap.com

The catalytic activity of DBH requires molecular oxygen and this compound. wikipedia.orgwikipedia.org this compound acts as an electron donor, reducing the cupric ions (Cu²⁺) at the active site of the enzyme to cuprous ions (Cu⁺). sci-hub.se This reduction allows the enzyme to be in its active state to facilitate the hydroxylation of dopamine. sci-hub.se In this process, this compound is oxidized to dehydroascorbic acid. wikipedia.orgsci-hub.se The reaction can be summarized as follows:

Dopamine + O₂ + this compound → Norepinephrine + H₂O + Dehydroascorbic acid wikipedia.org

This enzymatic reaction is unique as it is the only one in the synthesis of small-molecule neurotransmitters that is membrane-bound, occurring inside synaptic vesicles. wikipedia.org DBH is expressed in noradrenergic neurons in both the central and peripheral nervous systems, as well as in the chromaffin cells of the adrenal medulla. wikipedia.org Research has shown that a deficiency in this compound can lead to decreased concentrations of norepinephrine. mdpi.com Beyond its direct role as a cofactor for DBH, this compound also indirectly supports catecholamine synthesis by recycling tetrahydrobiopterin, a necessary cofactor for tyrosine hydroxylase, the rate-limiting enzyme in this pathway. mdpi.comresearchgate.net

Table 1: Role of this compound in Catecholamine Biosynthesis

Ten-Eleven Translocation (TET) Enzymes

This compound is a key cofactor for the Ten-Eleven Translocation (TET) family of enzymes (TET1, TET2, and TET3). mdpi.comnih.gov These enzymes are α-ketoglutarate-dependent dioxygenases that play a crucial role in epigenetic regulation by catalyzing the oxidation of 5-methylcytosine (5mC), a primary epigenetic mark, to 5-hydroxymethylcytosinet (5hmC). mdpi.comnih.govfrontiersin.org This process is the initial and rate-limiting step in active DNA demethylation. mdpi.com

The TET enzymes require Fe(II) and α-ketoglutarate as cofactors for their catalytic activity. nih.govfrontiersin.org this compound enhances TET activity by reducing the ferric iron (Fe³⁺) back to its ferrous state (Fe²⁺) at the catalytic center of the enzyme, thus regenerating the active form of the enzyme. ijbs.comresearchgate.net This function is crucial for maintaining the catalytic efficiency of TET proteins. acs.org The availability of this compound can, therefore, significantly influence the levels of 5hmC in the genome and modulate epigenetic control of gene activity. nih.gov

Studies have shown that this compound treatment can increase TET activity, leading to DNA demethylation and the reactivation of tumor suppressor genes in certain cancer cells. nih.gov The TET-mediated demethylation is essential for various biological processes, including cellular differentiation, embryonic development, and immune cell function. mdpi.comfrontiersin.org For instance, this compound promotes the differentiation of germinal center B cells to plasma cells by enhancing the catalytic activity of TET2 and TET3. frontiersin.orgbiorxiv.org

Table 2: this compound and TET Enzyme Function

Jumonji Histone Demethylases

This compound also serves as a cofactor for the Jumonji C (JmjC) domain-containing histone demethylases (JHDMs). mdpi.comnih.gov This large family of enzymes are also Fe(II) and α-ketoglutarate-dependent dioxygenases that are responsible for removing methyl groups from histone proteins. nih.govtandfonline.com Histone methylation is another critical epigenetic modification that influences chromatin structure and gene expression.

Similar to its role with TET enzymes, this compound is thought to promote the activity of JmjC histone demethylases by regenerating the catalytically active Fe²⁺ in the enzyme's active site. actanaturae.ru The JmjC domain is the catalytic core that carries out the demethylation of histones. nih.gov By acting as a cofactor, this compound facilitates the removal of various histone methyl marks, thereby playing a significant role in epigenetic regulation. nih.govtandfonline.com

The function of these histone demethylases is vital for a range of cellular processes, including somatic cell reprogramming and immune responses. nih.govresearchgate.net For example, this compound has been shown to induce the demethylation of H3K36me2/3 through the action of KDM2A/B, which are members of the JmjC family, thereby enhancing the efficiency of induced pluripotency. researchgate.net The modulation of histone demethylation by this compound highlights its importance in the intricate network of epigenetic control. nih.govactanaturae.ru

Table 3: this compound and Jumonji Histone Demethylase Function

Direct Scavenging of Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS)

This compound is a primary non-enzymatic antioxidant in plasma and tissues, effectively neutralizing a wide array of reactive oxygen species (ROS) and reactive nitrogen species (RNS). oregonstate.edubrieflands.compatsnap.com These harmful species are generated during normal metabolic processes, by immune cells, and through exposure to environmental toxins. oregonstate.edu The antioxidant activity of this compound stems from its ability to readily donate electrons, thereby neutralizing these reactive molecules. oregonstate.edupatsnap.com It is particularly effective against species such as the hydroxyl radical, superoxide radical, and hydrogen peroxide. frontiersin.orgresearchgate.net

A key antioxidant mechanism of this compound is its ability to donate a hydrogen atom to lipid radicals. brieflands.com This action is critical in halting the chain reactions of lipid peroxidation, a process that can cause significant damage to cell membranes. frontiersin.org By providing a hydrogen atom, this compound effectively neutralizes the lipid radical, preventing further propagation of oxidative damage.

This compound is an efficient quencher of singlet oxygen, a highly reactive form of oxygen. brieflands.comnih.gov It can deactivate singlet oxygen through a chemical quenching mechanism, which involves the oxidation of ascorbate. nih.govbmrat.orgmdpi.com The rate constant for the quenching of singlet oxygen by this compound has been measured, demonstrating its effectiveness in neutralizing this damaging species. researchgate.netkarger.com

This table presents the quenching rates of this compound on singlet oxygen at different pH levels, as determined by measuring the degradation of ascorbic acid in the presence of a photosensitizer under light storage. researchgate.net

The antioxidant mechanisms of this compound also include the removal of molecular oxygen. brieflands.com While the direct reaction with ground-state molecular oxygen (autoxidation) is slow in the absence of catalysts, this compound can react with oxygen, particularly in the presence of metal ions, to reduce its potential for generating harmful reactive species. nih.govtandfonline.com The degradation of this compound is significantly influenced by oxygen concentration, especially at elevated temperatures. nih.gov

When this compound donates a hydrogen atom or an electron to a free radical, it becomes the ascorbyl radical (also known as monodehydroascorbate). karger.comwikipedia.orgresearchgate.net This radical is relatively stable and less reactive than the radicals it neutralizes. wikipedia.orgresearchgate.net A key characteristic of the ascorbyl radical is that it preferentially reacts with other radicals rather than non-radical compounds, which helps to terminate free radical chain reactions. researchgate.net The ascorbyl radical can be reduced back to ascorbate by various cellular mechanisms or can disproportionate to form ascorbate and dehydroascorbic acid. encyclopedia.pubahajournals.org

Removal of Molecular Oxygen

Regeneration of Other Antioxidants (e.g., α-tocopherol)

This table shows the rapid rate at which this compound can donate a hydrogen atom to a tocopheroxyl radical, effectively regenerating α-tocopherol. brieflands.commdpi.com

Modulation of Intracellular Antioxidant Systems

This compound enhances the cellular antioxidant capacity not only by direct action but also by influencing the activity and levels of crucial antioxidant enzymes and molecules.

This compound has been shown to stimulate the activity of key antioxidant enzymes that are fundamental to cellular defense against oxidative damage. nih.govnih.gov These enzymes work in a coordinated fashion to neutralize harmful ROS.

Superoxide Dismutase (SOD): This enzyme catalyzes the dismutation of the superoxide radical (O₂⁻) into molecular oxygen (O₂) and hydrogen peroxide (H₂O₂). e-cep.org While some studies show no conclusive effect of this compound on SOD activity under normal physiological conditions, its influence becomes significant under conditions of oxidative stress. nih.gov For instance, in lymphocytes exposed to hydrogen peroxide, this compound supplementation led to increased baseline SOD activity. nih.gov Similarly, in the skin of mice exposed to gamma radiation, this compound pretreatment helped to prevent the radiation-induced decline in SOD activity. researchgate.net

Catalase (CAT): Catalase is responsible for the decomposition of hydrogen peroxide into water and oxygen, thus preventing the formation of the highly reactive hydroxyl radical. e-cep.org Research indicates that this compound can positively influence catalase activity. umk.pl In lymphocytes, supplementation with this compound resulted in elevated baseline catalase activity. nih.gov

Glutathione Peroxidase (GPx): This enzyme catalyzes the reduction of hydrogen peroxide and lipid hydroperoxides by utilizing reduced glutathione (GSH), thereby protecting cell membranes from lipid peroxidation. viamedica.ple-cep.org Studies have demonstrated that this compound supplementation can lead to a significant increase in the activity of glutathione peroxidase. researchgate.nettandfonline.com

Reduced glutathione (GSH) is a critical non-enzymatic antioxidant that plays a central role in cellular defense against oxidative stress. frontiersin.org this compound is intricately linked to the maintenance of GSH levels. The regeneration of this compound from its oxidized form, dehydroascorbic acid (DHA), can be mediated by GSH-dependent enzymes. mdpi.com

Research has shown that this compound supplementation can protect against the depletion of GSH levels induced by stressors like alcohol. tandfonline.com In a study on alcohol-induced hepatotoxicity in guinea pigs, this compound supplementation resulted in a faster restoration of GSH content compared to alcohol abstention alone. tandfonline.com Furthermore, increasing this compound levels in plants through genetic modification has been shown to lead to a corresponding increase in GSH levels, suggesting a coordinated regulation of these two vital antioxidants. pnas.org This interplay is crucial, as GSH is required for the activity of glutathione peroxidase and for the detoxification of xenobiotics. mdpi.comoup.com

Stimulation of Antioxidant Enzymes (e.g., Superoxide Dismutase, Catalase, Glutathione Peroxidase)

Regulation of ROS-Generating Enzymes (e.g., NADPH Oxidases, Xanthine Oxidase)

In addition to bolstering antioxidant defenses, this compound can also regulate the activity of enzymes that are major sources of cellular ROS.

NADPH Oxidases (NOXs): This family of enzymes generates superoxide by transferring electrons from NADPH to molecular oxygen and are key players in various physiological and pathological processes. smw.ch Excessive NOX activity can lead to oxidative stress. umk.pl this compound has been shown to down-regulate excessive NOX activity, helping to restore cellular redox homeostasis. umk.plencyclopedia.pub For example, this compound can inhibit the expression of the p47phox subunit of NADPH oxidase in microvascular endothelial cells. encyclopedia.pub

Xanthine Oxidase (XO): This enzyme catalyzes the oxidation of hypoxanthine and xanthine, producing uric acid and generating ROS such as superoxide and hydrogen peroxide in the process. nih.gov Supplementation with this compound has been found to offer significant protection against the hyperactivity of xanthine oxidase. umk.plencyclopedia.pub

Context-Dependent Pro-oxidant Activity in the Presence of Transition Metals

The pro-oxidant activity of this compound is most notably observed in the presence of free transition metal ions, particularly iron (Fe) and copper (Cu). nih.govtandfonline.com In this context, this compound acts as a reducing agent, converting ferric iron (Fe³⁺) to ferrous iron (Fe²⁺) and cupric copper (Cu²⁺) to cuprous copper (Cu¹⁺). nih.govscispace.com

This reduction of transition metals can then catalyze the formation of highly reactive oxygen species. The reduced iron (Fe²⁺) can react with hydrogen peroxide (H₂O₂) in the Fenton reaction to generate the highly damaging hydroxyl radical (•OH). nih.gov This pro-oxidant effect is dependent on several factors, including the redox state of the cellular environment, the availability of free transition metals, and the local concentration of this compound itself. researchgate.net

Induction of Apoptotic Cell Death at High Concentrations

At high, millimolar concentrations, this compound has been shown to selectively induce apoptotic cell death in various cancer cell lines, while leaving normal cells relatively unharmed. nih.govplos.org This cytotoxic effect is largely attributed to its pro-oxidant activity.

In the presence of transition metals, high concentrations of this compound can lead to the generation of hydrogen peroxide (H₂O₂). nih.govplos.org Cancer cells often have lower levels of antioxidant enzymes like catalase compared to normal cells, making them more susceptible to the damaging effects of H₂O₂ accumulation. ajol.info This buildup of H₂O₂ can induce oxidative stress, leading to DNA damage and ultimately triggering apoptosis. ajol.info

NF-κB/TNF-α Pathway Modulation

This compound has been shown to modulate the Nuclear Factor-kappa B (NF-κB) and Tumor Necrosis Factor-alpha (TNF-α) signaling pathway, a critical regulator of inflammation and immune responses. researchgate.net Under normal physiological conditions, NF-κB is kept in an inactive state in the cytoplasm through its association with an inhibitor protein. nih.gov Upon cellular stress, such as exposure to reactive oxygen species (ROS), this inhibitor is phosphorylated, leading to its degradation and the subsequent translocation of NF-κB to the nucleus. Once in the nucleus, NF-κB initiates the transcription of pro-inflammatory genes, including those for cytokines like TNF-α. nih.gov

This compound's antioxidant properties play a significant role in its modulation of this pathway. By reducing ROS levels, this compound can diminish a primary trigger for NF-κB activation. researchgate.net Research has demonstrated that this compound can suppress the activation of NF-κB induced by TNF-α. researchgate.net This inhibitory effect has been observed in various cell types and is associated with a decrease in the phosphorylation of the NF-κB inhibitor. researchgate.net For instance, in intestinal epithelial Caco-2 cells, treatment with an NF-κB inhibitor prevented the inhibitory effects of TNF-α on ascorbic acid uptake, highlighting the interplay between them. physiology.org

Furthermore, studies have indicated that this compound can influence the expression of pro-inflammatory cytokines. In mouse splenocytes, this compound has been found to downregulate the expression of TNF-α, along with other pro-inflammatory cytokines like IL-6 and IL-12, while upregulating anti-inflammatory cytokines. nih.gov Similarly, in brain tissue, this compound has been shown to significantly reduce TNF-α levels. nih.gov However, the effect of ascorbate on TNF-α levels can be context-dependent, as observed in the endometrial tissue of rats under high oxidative stress where it did not decrease TNF-α. nih.gov

The modulation of the NF-κB/TNF-α pathway by this compound is a crucial aspect of its anti-inflammatory and antioxidant functions. nih.gov By interfering with this signaling cascade, this compound helps to mitigate the inflammatory response that is often exacerbated by oxidative stress.

Nrf2, Ref-1, and AP-1 Activation

Beyond the NF-κB pathway, this compound also interacts with other critical transcription factors to bolster the cellular antioxidant defense system. These include Nuclear factor erythroid 2-related factor 2 (Nrf2), Redox effector factor-1 (Ref-1), and Activator protein-1 (AP-1). nih.govnih.gov

Nrf2 Activation:

Nrf2 is a key regulator of the antioxidant response element (ARE), which drives the expression of a wide array of antioxidant and detoxification enzymes. researchgate.net Under basal conditions, Nrf2 is kept in the cytoplasm by its inhibitor, Keap1. In the presence of oxidative or electrophilic stress, Nrf2 is released from Keap1 and translocates to the nucleus, where it binds to the ARE and initiates the transcription of protective genes. researchgate.net

This compound has been shown to promote the activity of Nrf2. nih.govnih.gov In hepatocytes, for example, this compound at concentrations of 1–10 µM can activate Nrf2, leading to the expression of antioxidant enzymes and a subsequent reduction in lipid hydroperoxides. nih.govresearchgate.net This activation of the Nrf2-ARE pathway is a significant mechanism by which this compound enhances the cell's intrinsic antioxidant capacity. researchgate.net However, it's noteworthy that very high concentrations of this compound (1 mM) have been suggested to potentially disrupt the Keap1/Nrf2/ARE pathway activation. nih.govresearchgate.net

Ref-1 and AP-1 Activation:

Ref-1 and AP-1 are also important transcription factors involved in the cellular response to stress. nih.gov Ref-1, a redox-sensitive protein, plays a role in the reduction and activation of other transcription factors, including AP-1. nih.gov AP-1 is a transcription factor that regulates gene expression in response to a variety of stimuli, including stress and growth factors.

The influence of this compound on Ref-1 and AP-1 appears to be more complex. Some studies suggest that only the oxidized form of ascorbate indirectly affects Ref-1 activity. nih.gov In the case of AP-1, this compound has been observed to have a silencing effect in certain cell types. For instance, in epidermal keratinocytes exposed to UV radiation, this compound supplementation led to reduced levels of the active component of AP-1, c-Jun. nih.gov A similar silencing of AP-1 activity by ascorbate was also noted in respiratory epithelial cells, resulting in lower levels of pro-inflammatory chemokines. nih.gov

The collective modulation of Nrf2, Ref-1, and AP-1 by this compound underscores its multifaceted role in orchestrating the cellular defense against oxidative stress. nih.gov By influencing these key transcription factors, this compound not only directly neutralizes ROS but also enhances the cell's own machinery for combating oxidative damage.

Data Tables

Table 1: Effects of this compound on NF-κB/TNF-α Pathway Components

Table 2: Effects of this compound on Nrf2, Ref-1, and AP-1

L-Galactose Pathway (Smirnoff-Wheeler pathway) in Plants

The primary and most well-understood route for this compound biosynthesis in plants is the L-galactose pathway, also known as the Smirnoff-Wheeler pathway. mdpi.commba.ac.uk This pathway begins with D-glucose-6-phosphate and proceeds through a series of enzymatic reactions to produce this compound. mdpi.com

Several key enzymes and their corresponding genes are crucial for the L-galactose pathway. frontiersin.orgmdpi.com These enzymes catalyze the sequential conversion of intermediates, ultimately leading to the synthesis of this compound. frontiersin.orgmdpi.com The major enzymes in this pathway are GDP-D-mannose pyrophosphorylase (GMP), GDP-L-galactose phosphorylase (GGP), and L-galactono-1,4-lactone dehydrogenase (GLDH). frontiersin.orgmdpi.commdpi.com

The biosynthesis of this compound in plants is influenced by various environmental cues, with light and temperature being significant factors. researchgate.net Light, in particular, stimulates the production of this compound. researchgate.netfrontiersin.org This response is linked to the increased need for antioxidants to counteract the reactive oxygen species generated during photosynthesis. frontiersin.org The expression of key biosynthetic genes, such as GGP and GLDH, is induced by high light conditions. frontiersin.org Conversely, darkness leads to a reduction in this compound levels, partly due to the degradation of the GMP enzyme. frontiersin.org

The final step in the L-galactose pathway, the oxidation of L-galactono-1,4-lactone to this compound, is catalyzed by L-galactono-1,4-lactone dehydrogenase (GLDH). frontiersin.org This enzyme is located on the inner mitochondrial membrane and is directly linked to the mitochondrial electron transport chain. mdpi.comnih.gov GLDH donates electrons to cytochrome c, positioning this biosynthetic step between complexes III and IV of the respiratory chain. frontiersin.orgnih.gov This connection highlights a direct interplay between this compound synthesis and mitochondrial respiration. uantwerpen.be

Regulation by Environmental Factors (e.g., Light, Temperature)

Inability of Humans and Other Mammals to Synthesize this compound (Due to GULO gene mutation)

Unlike most plants and many animals, humans, other primates, guinea pigs, and some bat species are unable to synthesize their own this compound. taylorandfrancis.comnih.govmdpi.comquora.com This inability is due to a series of mutations in the gene encoding L-gulono-γ-lactone oxidase (GULO). taylorandfrancis.comnih.govgeneticeducation.co.in This enzyme catalyzes the final step in the animal biosynthesis pathway, the conversion of L-gulono-γ-lactone to this compound. geneticeducation.co.inontosight.ai The human GULO gene, located on chromosome 8, has become a non-functional pseudogene over the course of evolution, necessitating the intake of this compound through diet. nih.govgeneticeducation.co.in

Table of Compounds

Conversion of Dehydroascorbate to this compound

The oxidation of this compound yields the short-lived monodehydroascorbate (MDHA) radical. nih.govoup.com MDHA can be directly reduced back to ascorbate or can undergo non-enzymatic disproportionation, where two MDHA molecules react to form one molecule of ascorbate and one molecule of dehydroascorbate (DHA). nih.govresearchgate.netmdpi.com

DHA, the two-electron oxidized form of ascorbate, can then be recycled back to this compound. researchgate.netbiorxiv.org This reduction is a critical step, as DHA will otherwise undergo irreversible hydrolysis to form 2,3-diketogulonic acid, resulting in a loss of the ascorbate pool. researchgate.netoup.comresearchgate.net The regeneration of ascorbate from DHA is primarily an enzymatic process. researchgate.net

Enzymes Involved in Recycling (e.g., Monodehydroascorbate Reductase, Dehydroascorbate Reductase)

Two key enzymes are responsible for the recycling of this compound: monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR). nih.govresearchgate.net These enzymes are central to the ascorbate-glutathione cycle, which plays a significant role in cellular antioxidant defense. mdpi.comoup.com

Monodehydroascorbate Reductase (MDHAR): This enzyme catalyzes the reduction of the monodehydroascorbate (MDHA) radical back to this compound. nih.govresearchgate.net MDHAR utilizes NAD(P)H as an electron donor for this reaction. researchgate.netoup.com The direct reduction of MDHA by MDHAR is an efficient way to regenerate ascorbate before it can disproportionate into DHA. oup.commdpi.com

Dehydroascorbate Reductase (DHAR): This enzyme facilitates the reduction of dehydroascorbate (DHA) to this compound. nih.govresearchgate.netbiorxiv.org DHAR uses glutathione (GSH) as the reducing agent, which is in turn oxidized to glutathione disulfide (GSSG). researchgate.netoup.com The oxidized glutathione is then regenerated by glutathione reductase (GR) using NADPH as a reductant. researchgate.netresearchgate.net The action of DHAR is vital to prevent the irreversible degradation of DHA. nih.govoup.com

Table 1: Key Enzymes in this compound Recycling

Sodium-Dependent Vitamin C Transporters (SVCT1 and SVCT2)

The primary mechanism for the uptake of the reduced form of vitamin C, L-ascorbic acid, is through the action of two sodium-dependent vitamin C co-transporters, SVCT1 and SVCT2. nih.govresearchgate.net These transporters are members of the solute carrier family 23 (SLC23). frontiersin.org SVCT1 and SVCT2 are surface glycoproteins encoded by two different genes, SLC23A1 and SLC23A2 respectively, and share approximately 65% of their amino acid sequence. scilit.commdpi.com Despite their structural similarities, they exhibit distinct tissue distributions and functional characteristics that point to different physiological roles. scilit.comnih.govresearchgate.net

SVCT1 and SVCT2 are responsible for the active transport of this compound into cells, a process that allows for the accumulation of the vitamin to concentrations significantly higher than those in the extracellular fluid. frontiersin.orgmdpi.commdpi.com This transport is an energy-dependent process that moves this compound against its concentration gradient. ontosight.ai The energy required for this "uphill" transport is derived from the electrochemical gradient of sodium ions (Na+), which is maintained by the Na+/K+-ATPase pump. frontiersin.orgmdpi.com

The stoichiometry of this co-transport process has been determined to be two sodium ions for every one molecule of this compound transported into the cell. frontiersin.orgmdpi.comtandfonline.comphysiology.org This 2:1 ratio of Na+ to ascorbate demonstrates the secondary active transport mechanism that drives the high level of vitamin C accumulation in cells. mdpi.com The transport process is also sensitive to temperature and pH, with an optimal pH of approximately 7.5. frontiersin.orgtandfonline.com Furthermore, the activity of SVCT2 has been shown to be dependent on the presence of calcium (Ca2+) and magnesium (Mg2+) ions. frontiersin.org

SVCT1 and SVCT2 exhibit distinct patterns of tissue expression and different affinities for this compound, which reflects their specialized roles in vitamin C homeostasis. scilit.comnih.govresearchgate.net SVCT1 is predominantly found in epithelial tissues, where it is responsible for the bulk transport and whole-body homeostasis of vitamin C. researchgate.netmdpi.comoup.comoup.com In contrast, SVCT2 is more widely distributed and is crucial for protecting metabolically active cells from oxidative stress by ensuring they have an adequate supply of ascorbate. scilit.comnih.govresearchgate.net

SVCT1 is characterized as a low-affinity, high-capacity transporter. mdpi.commdpi.com Its expression is largely confined to the epithelial cells of the intestine, where it mediates the absorption of dietary vitamin C, and the kidneys, where it is responsible for the reabsorption of ascorbate from the glomerular filtrate. frontiersin.orgresearchgate.netmdpi.commdpi.com It is also found in the liver. researchgate.net The lower affinity of SVCT1 is well-suited for handling the higher concentrations of ascorbate found in the diet and the kidney tubules. frontiersin.org

SVCT2 , on the other hand, is a high-affinity, low-capacity transporter. frontiersin.orgmdpi.com It is expressed in a wide variety of tissues, including the brain, eyes, placenta, and neuroendocrine, exocrine, and endothelial tissues. researchgate.netplos.org This high-affinity transporter enables cells in these tissues to take up ascorbate from the low micromolar concentrations typically found in the blood and accumulate it to millimolar levels internally. frontiersin.orgmdpi.com This is particularly critical in the brain, where high concentrations of ascorbate are essential for neuronal function and protection. frontiersin.orgmdpi.com

Table 1: Comparison of SVCT1 and SVCT2 Characteristics

The expression and activity of SVCT1 and SVCT2 are subject to regulation at multiple levels, including transcriptional, translational, and post-translational modifications, ensuring a fine-tuned control of vitamin C uptake. scilit.comfrontiersin.orgthomashealthblog.com

Transcriptional Regulation: The transcription of the SLC23A1 and SLC23A2 genes can be influenced by various factors. For instance, studies have shown that the expression of SVCT1 and SVCT2 mRNA can be upregulated in response to vitamin C deficiency in certain tissues, representing an adaptive response to increase vitamin C uptake. mdpi.com The transcription factors Hepatocyte Nuclear Factor 1 alpha (HNF1α) and Specificity Protein 1 (Sp1) are known to regulate the promoter activity of SLC23A1 (SVCT1) and SLC23A2 (SVCT2), respectively. mdpi.com Hormones and other signaling molecules also play a role; for example, glucocorticoids have been shown to increase SVCT2 mRNA levels in osteoblastic cells. thomashealthblog.com In contrast, high concentrations of ascorbate have been observed to downregulate SVCT1 expression in human colon carcinoma cells. frontiersin.org

Post-translational Modification: Both SVCT1 and SVCT2 proteins have potential sites for post-translational modifications such as glycosylation and phosphorylation, which can regulate their activity. frontiersin.orgmdpi.comtandfonline.com Phosphorylation by protein kinase C (PKC) has different effects on the two transporters. In the case of SVCT1, PKC activation leads to a decrease in its translocation from the cytoplasm to the cell membrane. mdpi.com For SVCT2, phosphorylation by PKC is thought to induce a conformational change in the transporter. mdpi.com Additionally, SVCT1 has a potential phosphorylation site for protein kinase A (PKA). tandfonline.com The translocation of SVCT transporters from intracellular compartments to the cell membrane is another important regulatory mechanism that can rapidly increase ascorbate uptake without altering gene expression. frontiersin.org

A naturally occurring short isoform of SVCT2, resulting from alternative splicing, has been identified and is unable to transport ascorbate. frontiersin.org This isoform can negatively regulate the function of the full-length SVCT2 and can also partially inhibit SVCT1, likely through the formation of hetero-oligomers. frontiersin.org

Tissue-Specific Expression and Affinity Characteristics

Volume-Sensitive Anion Channels (VSOAC)

Volume-sensitive anion channels (VSOACs), also known as volume-regulated anion channels (VRACs), are a key pathway for ascorbate efflux, particularly in response to osmotic stress. medsci.orgresearchgate.net These channels are activated when a cell swells due to an increase in intracellular water. medsci.orgfrontiersin.org The resulting channel opening allows for the passage of chloride and organic osmolytes, including ascorbate, out of the cell. medsci.orgresearchgate.net This efflux helps to restore normal cell volume, a process termed regulatory volume decrease. researchgate.net In the central nervous system, VSOACs in astrocytes have been shown to mediate the release of ascorbate in response to cell swelling. medsci.org The LRRC8 family of proteins are essential components of these channels. researchgate.netuniprot.org

Ca²⁺-Dependent Anion Channels

The release of this compound from cells can also be mediated by anion channels that are activated by an increase in intracellular calcium (Ca²⁺) concentration. researchgate.netnih.govtermedia.pl Research on coronary artery endothelial cells has demonstrated that agonists like ATP, which elevate intracellular Ca²⁺, can potentiate the release of ascorbate. nih.gov This Ca²⁺-mediated efflux was found to be dependent on the presence of extracellular chloride and was inhibited by blockers of anion channels, suggesting that a Ca²⁺-dependent anion channel is responsible for the transport. nih.govoup.com

Gap-Junction Hemichannels

Gap junctions are intercellular channels formed by proteins called connexins, which allow direct communication between adjacent cells. nih.govoup.com A gap junction is composed of two hemichannels, or connexons, one from each cell. nih.gov Under certain physiological or pathological conditions, these hemichannels can exist in an un-docked state on the cell surface and function as a conduit between the cytoplasm and the extracellular environment. oup.comvaincrealzheimer.org The opening of connexin hemichannels, such as those formed by Connexin 26, Connexin 32, and Connexin 43, has been shown to permit the release of small molecules, including ATP, glutathione, and this compound. oup.comtandfonline.comresearchgate.net

Exocytosis of Secretory Vesicles

Exocytosis is a process where cells release substances stored in intracellular vesicles by fusing the vesicular membrane with the plasma membrane. wikipedia.org This mechanism is utilized for ascorbate efflux in specialized secretory cells, such as the chromaffin cells of the adrenal medulla and neuroendocrine cells. researchgate.netpnas.org These cells store high concentrations of this compound within secretory vesicles, alongside hormones and neurotransmitters like catecholamines. pnas.org this compound also functions as an essential cofactor for the enzyme dopamine β-monooxygenase within these vesicles, which is necessary for the synthesis of norepinephrine from dopamine. wikipedia.org Upon receiving a specific stimulus, these vesicles undergo exocytosis, releasing their entire contents, including ascorbate, into the extracellular space. researchgate.netwikipedia.org

Homo- and Hetero-exchange Systems

This compound can be released from cells via membrane transporters that facilitate its exchange for other molecules. researchgate.netnih.govfrontiersin.org A prominent example of this is the ascorbate-glutamate hetero-exchange system, which is particularly active in glial cells (astrocytes) of the brain. brieflands.commdpi.com This process involves specific transporters that mediate the efflux of intracellular ascorbate in exchange for the uptake of extracellular glutamate. mdpi.com This hetero-exchange is believed to play a neuroprotective role by helping to clear excess glutamate from the synaptic space, thereby protecting neurons from excitotoxicity. brieflands.commdpi.com In addition to hetero-exchange, ascorbate-ascorbate homo-exchange has also been proposed as a potential efflux mechanism. termedia.pl

Mitochondrial Transport of this compound

The presence of this compound in mitochondria is critical for protecting these organelles from oxidative damage generated as a byproduct of cellular respiration. researchgate.net In vivo studies have confirmed that mitochondria contain this compound, and its concentration within these organelles can be increased through dietary supplementation. researchgate.netnih.gov The transport of vitamin C into the mitochondrial matrix is a complex process, with several proposed mechanisms involving different transporters and forms of the vitamin. The primary debate centers on whether the vitamin enters in its reduced form (this compound) or its oxidized form, dehydroascorbic acid (DHA). researchgate.netmdpi.com

Transport via Glucose Transporters (GLUTs)

A significant body of evidence suggests that mitochondria import vitamin C primarily in its oxidized form, dehydroascorbic acid (DHA), via glucose transporters (GLUTs). nih.govoup.com Mammalian mitochondria themselves are generally considered unable to transport the reduced this compound directly. oup.com Instead, DHA is transported into the mitochondrial matrix by facilitated diffusion through specific GLUT isoforms that are localized to the mitochondrial membranes. mdpi.commdpi.com

GLUT1: This transporter has been identified on the inner mitochondrial membrane and is proposed to facilitate the uptake of DHA. researchgate.netoup.com Studies in human kidney cells support the role of GLUT1 in transporting DHA into mitochondria. nih.gov

GLUT10: Research has identified GLUT10 as a key mitochondrial DHA transporter, particularly in smooth muscle cells and insulin-stimulated adipocytes. oup.com The localization of GLUT10 to mitochondria allows it to mediate the import of DHA, which subsequently increases the intramitochondrial concentration of this compound after its reduction, protecting cells from oxidative injury. oup.com

Once inside the mitochondrial matrix, DHA is rapidly and efficiently reduced back to this compound. researchgate.netnih.gov This intracellular recycling is crucial because it effectively traps the vitamin within the organelle and regenerates its antioxidant capacity. core.ac.uknih.gov

Transport via Sodium-Dependent Vitamin C Transporter 2 (SVCT2)

Another proposed mechanism involves the direct transport of the reduced form, this compound, by the sodium-dependent vitamin C transporter 2 (SVCT2). nih.govresearchgate.net Research has provided evidence for the localization of SVCT2 to mitochondria in certain cell types.

In human HEK-293 cells, SVCT2 immunoreactivity was predominantly associated with mitochondria. nih.gov

Immunoblotting of highly purified mitochondrial fractions from these cells confirmed the presence of the SVCT2 protein. nih.gov

Silencing the expression of SVCT2 with siRNA resulted in a significant decrease in both the mitochondrial SVCT2 protein and the rate of mitochondrial this compound transport. nih.gov

However, the function of SVCT2 in the mitochondrial environment is thought to be different from its high-affinity function at the plasma membrane. nih.gov The mitochondrial intermembrane space has a low sodium and high potassium concentration, which causes mitochondrial SVCT2 to function as a low-affinity transporter. nih.govd-nb.info The role of mitochondrial SVCT2 remains a subject of debate, as some studies have failed to detect its colocalization with mitochondria in other cell types, such as rat neuronal cultures, suggesting that its role may be tissue-specific. mdpi.com

The Mitochondrial Ascorbate Transporter (MAT)

Further research has led to the identification and characterization of a novel mitochondrial ascorbate transporter (MAT) from rat liver and potato mitochondria. frontiersin.org This transporter is distinct from SVCT2 and GLUTs. frontiersin.org Key characteristics of MAT include:

It is sodium-independent. frontiersin.org

It has a molecular mass in the range of 28–35 kDa. frontiersin.org

It is stimulated by a proton gradient, suggesting it functions as an ascorbate-proton symporter. frontiersin.org

Mitochondrial Recycling of this compound

Regardless of the uptake mechanism, the reduction of DHA to this compound within the mitochondrial matrix is a critical step for its accumulation and function. core.ac.uk This recycling process is dependent on several intramitochondrial systems:

Glutathione (GSH): DHA reduction depends heavily on reduced glutathione. researchgate.net

Thioredoxin Reductase: The thioredoxin system may also contribute to the regeneration of this compound from DHA. researchgate.net

Electron Transport Chain: The mitochondrial electron transport chain itself can provide electrons for DHA reduction, with evidence pointing to a site on complex III as being involved in the recycling process. researchgate.netnih.govresearchgate.net

This rapid internal reduction ensures that this compound is available to scavenge reactive oxygen species at their primary site of production, thereby protecting the mitochondria from oxidative damage. nih.gov

Data Table: Mitochondrial Transporters of this compound and its Derivatives

Preferential Stimulation of Collagen-Specific mRNA

A primary and well-documented effect of this compound is its ability to selectively increase the synthesis of collagen proteins. mdpi.comnih.gov This stimulation is closely linked to a notable increase in the messenger RNA (mRNA) levels of procollagen genes, particularly for type I and type III collagen. mdpi.comresearchgate.net Research indicates that this compound can enhance the transcription of collagen genes and also stabilize the collagen mRNA, leading to increased protein synthesis for tissue repair. oregonstate.eduresearchgate.net This effect appears to be specific, as the synthesis of non-collagenous proteins is not similarly increased. mdpi.com

One proposed mechanism for this specific action involves a feedback loop where the accumulation of procollagen within the cell inhibits its own synthesis. By acting as a cofactor for prolyl and lysyl hydroxylases, this compound promotes the hydroxylation and subsequent secretion of procollagen, thereby releasing this translational inhibition and allowing for increased collagen gene transcription. mdpi.com Another hypothesis suggests that lipid peroxidation, which can be induced by this compound, may mediate the increase in collagen gene expression. researchgate.net Studies have shown that malondialdehyde, a product of lipid peroxidation, can increase collagen gene expression. researchgate.net

Other Effects on Transcriptional and Translational Levels

Beyond its pronounced effect on collagen, this compound influences a broader range of gene expression profiles. In some contexts, it can inhibit the expression of genes involved in cell cycle progression, such as tRNA synthetases and translation initiation factor subunits, leading to an antiproliferative effect. plos.org Conversely, it has been shown to upregulate the expression of tumor suppressor genes like p53 while inhibiting genes that promote tumor development. oatext.com

The molecule's influence can also be context-dependent, showing moderate effects on gene expression under normal physiological conditions but a significant modulatory effect upon an inflammatory stimulus. cambridge.orgnih.gov For instance, upon inflammatory activation, this compound supplementation has been linked to differential activation of the MyD88-dependent signaling pathway, which activates key transcription factors like NF-κB and AP1. cambridge.org In malignant T-lymphocytes, high doses of ascorbic acid have been observed to inhibit the activity and protein expression of matrix metalloproteinase-2 (MMP-2), a key enzyme in cancer metastasis, without affecting its transcriptional levels. spandidos-publications.com

Influence on Histone Demethylases

In addition to its role in DNA demethylation, this compound is also a cofactor for Jumonji C (JmjC) domain-containing histone demethylases (JHDMs). nih.govnih.govcybermedlife.eu These enzymes are responsible for removing methyl groups from histone proteins, which are key components of chromatin. cybermedlife.eu Histone methylation is a critical epigenetic modification that can either activate or repress gene transcription depending on the specific site and degree of methylation.

This compound is required for the optimal activity of JHDMs, facilitating the removal of repressive histone marks such as H3K9 and H3K27 methylation. cybermedlife.eufrontiersin.org By promoting the activity of these demethylases, this compound can help to erase epigenetic barriers, for example, during the reprogramming of somatic cells into induced pluripotent stem cells (iPSCs). benthamdirect.comfrontiersin.org This function highlights this compound's broad impact on chromatin architecture and the regulation of gene expression. nih.govcybermedlife.eu

Stimulation of Leukocyte Production and Function (Neutrophils, Lymphocytes, Phagocytes)

This compound has been demonstrated to stimulate the production and function of various leukocytes, which are essential for identifying and eliminating pathogens. oregonstate.edu It enhances the proliferation and differentiation of B- and T-lymphocytes, key components of the adaptive immune response. oregonstate.educonsensus.appnih.gov Studies have shown that this compound is necessary for the maturation of T-cells and can increase the production of B-lymphocytes, which are responsible for producing antibodies. consensus.appdoctorsformulas.com Furthermore, it supports the function of phagocytes, such as neutrophils and macrophages, which engulf and digest foreign particles. oregonstate.edumedicalresearchjournal.org

Enhancement of Cellular Motility, Chemotaxis, and Phagocytosis

This compound plays a significant role in the directed movement and activity of immune cells. It has been shown to enhance cellular motility and chemotaxis, the process by which immune cells are attracted to a site of infection or inflammation. oregonstate.edunih.govakjournals.com Specifically, this compound improves the migration of neutrophils to infection sites. nih.govmedicalresearchjournal.org Once at the site, it enhances their phagocytic capacity, which is the engulfment of microorganisms and cellular debris. oregonstate.edunih.govmedicalresearchjournal.org

Protection of Immune Cells from Oxidative Damage

Immune cells, particularly phagocytes like neutrophils, generate reactive oxygen species (ROS) to kill pathogens. oregonstate.edu While essential for microbial killing, these ROS can also cause oxidative damage to the immune cells themselves. oregonstate.edu this compound, being a potent antioxidant, accumulates in high concentrations within these cells, protecting them from this self-inflicted oxidative damage. oregonstate.edunih.govakjournals.com It neutralizes ROS, thereby preserving the integrity and function of leukocytes. nih.govclinisciences.com This protective role is crucial for maintaining a robust and effective immune response.

Reduction of Ferric (Fe3+) to Ferrous (Fe2+) Ions

One of the key chemical properties of this compound is its ability to act as a reducing agent, meaning it readily donates electrons. oregonstate.educlinisciences.com In the context of iron metabolism, this compound facilitates the reduction of ferric iron (Fe3+) to ferrous iron (Fe2+). clinisciences.comnih.govresearchgate.net This conversion is critical because ferrous iron is more soluble and readily absorbed by the body. nih.govmdpi.com The reduction of Fe3+ to Fe2+ by ascorbate can occur through the formation of an intermediate chelate complex. nih.govmdpi.com

Promotion of Dietary Non-Heme Iron Absorption

Dietary iron exists in two forms: heme iron, derived from hemoglobin and myoglobin in animal tissues, and non-heme iron, found in plant-based foods and iron supplements. nih.gov The absorption of non-heme iron is significantly influenced by other dietary components. nih.gov this compound is a powerful enhancer of non-heme iron absorption. nih.govacs.orgscispace.com By reducing ferric iron to the more soluble ferrous form, this compound increases its bioavailability for absorption in the small intestine. researchgate.netnih.govacs.org It forms a soluble chelate with iron at the acidic pH of the stomach, which remains soluble at the more alkaline pH of the duodenum, further facilitating its uptake. nih.govscispace.com The enhancing effect of this compound on non-heme iron absorption is directly proportional to the amount of ascorbic acid present in a meal. nih.gov

Interactive Data Table: this compound's Role in Biological Systems

Stabilization of Iron-Binding Proteins

This compound plays a significant role in iron metabolism, primarily by enhancing the absorption of non-heme iron from the gastrointestinal tract. scispace.comresearchgate.net It achieves this by reducing ferric iron (Fe³⁺) to ferrous iron (Fe²⁺), the form more readily absorbed by the body. scispace.comresearchgate.net This reduction is critical for the effective transport of iron into cells.

Beyond absorption, this compound is involved in the stabilization of iron-binding proteins. scispace.comresearchgate.net It has the capacity to form chelate complexes with both ferrous (Fe²⁺) and ferric (Fe³⁺) ions. mdpi.comnih.gov The alpha oxo-hydroxy and di-hydroxy groups within the ascorbic acid structure are particularly suited for binding with iron. nih.gov This interaction is important for the normal function of proteins involved in iron metabolism, such as transferrin and ferritin. nih.gov By binding to iron, ascorbic acid can influence its uptake, transport, and storage within the body. nih.gov While it is a powerful antioxidant, in the presence of excess iron, this compound can act as a pro-oxidant by reducing Fe³⁺ to Fe²⁺, which can then participate in reactions that generate free radicals. nih.gov

High-Performance Liquid Chromatography (HPLC)

High-Performance Liquid Chromatography (HPLC) is a predominant technique for the analysis of this compound due to its high selectivity, specificity, and rapidity. wisdomlib.orgasianpubs.org The method separates components of a mixture for subsequent identification and quantification. scribd.com Reversed-phase HPLC (RP-HPLC) is commonly employed, utilizing a non-polar stationary phase and a polar mobile phase. scribd.comuin-alauddin.ac.id This setup allows for the effective separation of polar compounds like this compound. helixchrom.com

The most common method for detecting this compound after HPLC separation is through ultraviolet (UV) absorption. asianpubs.org this compound exhibits a strong absorbance in the UV spectrum, with detection wavelengths typically set between 243 nm and 266 nm for optimal sensitivity. asianpubs.orgasianpubs.orgoiv.int The selection of the mobile phase, often an acidic buffer mixed with an organic solvent like methanol, is critical for achieving good separation and a stable baseline. asianpubs.orgasianpubs.org For instance, a mobile phase consisting of HPLC grade water adjusted to pH 2.2 with sulfuric acid and methanol (80:20) has been successfully used with UV detection at 243 nm. asianpubs.org Another method for analyzing wine samples utilizes a reversed-phase column with UV detection at 266 nm. oiv.intoiv.int The simplicity and robustness of HPLC-UV make it suitable for routine analysis in many quality control laboratories. asianpubs.org

Table 1: Examples of HPLC-UV Methods for this compound Determination

For enhanced sensitivity, HPLC systems can be integrated with an electrochemical detector (ECD). wisdomlib.orgmdpi.com HPLC-ECD has proven to be a highly selective and sensitive technique for evaluating this compound in food and biological fluids. mdpi.com This method relies on the electroactive nature of this compound, which can be readily oxidized at the surface of an electrode. scispace.com The integration of amperometric or coulometric detectors with HPLC significantly improves detection limits. wisdomlib.orgnih.gov

A developed HPLC-ECD method demonstrated superior sensitivity, simpler operation, and more rapid detection compared to HPLC with diode-array detection (DAD). mdpi.comnih.gov The validation of one such method showed a limit of detection (LOD) as low as 0.0043 µg/mL and a linear range of 0.1 to 20 µg/mL, making it suitable for samples with very low this compound content. mdpi.comnih.govresearchgate.net The accuracy of the HPLC-ECD method has been reported in the range of 96.90% to 102.00%. mdpi.com

Table 2: Performance Comparison of an HPLC-ECD Method

UV Detection

Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS)

Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS) is a highly advanced analytical method that provides exceptional sensitivity and specificity for the quantification of this compound. scielo.brscielo.brresearchgate.net This technique couples the high-resolution separation power of UPLC with the precise detection and structural confirmation capabilities of tandem mass spectrometry. nih.gov It is considered a gold standard in chemical detection and is particularly valuable for analyzing this compound in complex biological matrices such as milk and human plasma, where concentrations can be very low and interference from other components is high. scielo.brresearchgate.netnih.gov

One validated UPLC-MS/MS method for determining this compound in milk samples reported a limit of detection (LOD) of 1.5 µg L⁻¹ and a limit of quantification (LOQ) of 5.0 µg L⁻¹. scielo.br The method demonstrated excellent linearity and high recovery rates (>90%), making it fast, accurate, and reproducible. scielo.brresearchgate.net The chromatographic separation is typically performed on a sub-2 µm particle column, such as a Waters Acquity UPLC® BEH C18, with an isocratic mobile phase, leading to very short analysis times. scielo.br

Table 3: UPLC-MS/MS Method Parameters for this compound Analysis in Milk

High-Performance Thin Layer Chromatography (HPTLC)

High-Performance Thin Layer Chromatography (HPTLC) offers a cost-effective, rapid, and high-throughput alternative for the quantitative analysis of this compound. mdpi.comsepscience.combibliomed.org This planar chromatography technique allows for the parallel analysis of multiple samples on a single plate, which significantly reduces the analysis time per sample. sigmaaldrich.com HPTLC is particularly advantageous due to its minimal sample clean-up requirements and high matrix tolerance, making it well-suited for complex samples like fruit juices and herbal extracts. sepscience.combibliomed.orgsigmaaldrich.com

In HPTLC, separation occurs on a plate coated with a layer of adsorbent, typically silica gel 60 F254. bibliomed.orgresearchgate.net The plate is then developed in a chamber containing a suitable mobile phase. For this compound, a greener HPTLC method was developed using a mobile phase of water-ethanol (70:30 v/v). mdpi.com Quantitative analysis is performed by scanning the plate with a densitometer at a specific wavelength, such as 254 nm or 265 nm, where this compound absorbs light. mdpi.comresearchgate.net The method can be validated according to ICH guidelines for linearity, accuracy, and precision. bibliomed.org

Table 4: Research Findings for HPTLC Analysis of this compound

Amperometric Methods

Amperometric detection is a prominent electrochemical technique for this compound analysis, leveraging its electroactive nature. electrochemsci.org This method involves the application of a constant potential to a working electrode and measuring the resulting current, which is proportional to the concentration of the analyte being oxidized or reduced. This compound is readily oxidized, making it an ideal candidate for amperometric determination. researchgate.net

Researchers have developed various amperometric sensors and systems to enhance the selectivity and sensitivity of this compound detection. These often involve the modification of electrode surfaces with materials that facilitate the electrochemical reaction. For instance, a glassy carbon electrode modified with octacyanomolybdate-doped-poly(4-vinylpyridine) has been successfully used for the direct estimation of this compound in orange juice and pharmaceutical tablets. researchgate.net Another approach involves flow-injection analysis (FIA) systems, where a sample is injected into a carrier stream that transports it to the amperometric detector. nih.gov An FIA method based on the photochemical reduction of Methylene Blue has been reported for the assay of this compound, with a detection limit of 1.9 µg ml-1. nih.gov

Amperometric biosensors, which integrate a biological recognition element with an electrochemical transducer, offer high specificity. electrochemsci.orgrecentonline.ro For example, an amperometric biosensor for this compound has been developed by immobilizing ascorbate oxidase onto a nylon net. redalyc.org The enzyme specifically catalyzes the oxidation of this compound, and the resulting change in oxygen concentration is monitored by an oxygen electrode. redalyc.org

Table 1: Comparison of Amperometric Methods for this compound Determination

Coulometric Methods

Coulometry is an absolute analytical method based on Faraday's law of electrolysis, where the total charge passed during an electrochemical reaction is measured. This technique offers the advantage of not requiring calibration standards, provided the reaction proceeds with 100% current efficiency. core.ac.uk For this compound, coulometric titration with electrogenerated iodine is a common approach. acs.orgscribd.com In this method, iodide is electrochemically oxidized to iodine, which then reacts with this compound. The endpoint is typically detected using a starch indicator. acs.org

Controlled-potential coulometry has also been applied to the determination of this compound, particularly in paper-based electrochemical devices. core.ac.uk This method involves applying a specific potential to electrolyze the entire sample and integrating the resulting current over time to obtain the charge. core.ac.uk A study utilizing a paper-based carbon working electrode demonstrated a limit of detection of 40 µM for ascorbic acid in fruit juices. core.ac.uk

Furthermore, coulometric detectors can be coupled with high-performance liquid chromatography (HPLC) to provide sensitive and selective analysis of electroactive species like this compound. nih.gov A comparison between amperometric and coulometric detectors revealed that the coulometric detector was significantly more sensitive for ascorbic acid detection. nih.gov

Table 2: Coulometric Methods for this compound Analysis

Voltammetric Techniques (Square Wave, Differential Pulse, Cyclic Voltammetry)

Voltammetric techniques are powerful electrochemical methods that involve applying a varying potential to an electrode and measuring the resulting current. nih.gov These methods are widely used for this compound determination due to their rapidity, simplicity, and low detection limits. jscimedcentral.com

Cyclic Voltammetry (CV) is often used to study the electrochemical behavior of this compound and to investigate the properties of modified electrodes. recentonline.rojscimedcentral.com It provides information about the oxidation and reduction processes of the analyte. nih.gov

Differential Pulse Voltammetry (DPV) and Square Wave Voltammetry (SWV) are more sensitive techniques used for quantitative analysis. recentonline.rojscimedcentral.com They employ potential waveforms that minimize background currents, resulting in enhanced signal-to-noise ratios.

Numerous studies have reported the use of various modified electrodes to improve the sensitivity and selectivity of voltammetric methods for this compound. For example, a glassy carbon electrode modified with a multi-walled carbon nanotube-tetradecyltrimethylammonium bromide film has been used to study the electro-oxidation of ascorbic acid using DPV, CV, and SWV. jscimedcentral.com Another study utilized a carbon paste electrode modified with 2,7-bis (ferrocenylethynyl) fluoren-9-one for the electrocatalytic determination of ascorbic acid using CV and DPV. jscimedcentral.com These modified electrodes often exhibit electrocatalytic activity towards the oxidation of ascorbic acid, allowing for determination at lower potentials and with reduced interference from other substances. encyclopedia.pub

Table 3: Voltammetric Techniques for this compound Determination