Pantothenic Acid, Biotin, Vit B6, Folate, and Vit B12

Pantothenate Acid

Ionized form of pantothenic acid

Pantothenic acid

• precursor of Coenzyme A (CoA)

• Biosynthesis of CoA uses pantothenate, ATP, and cysteine as substrates

• There are 5 enzymatic steps in CoA biosynthesis

  • Pantothenate —> pantothenate kinase ( along with Mg2+ and ATP to ADP)—> 4’ phosphopantothenate

  • This is also the rate-limiting steps

Pantothenic Acid Sources

• virtually all foods

  • excellent sources: animal organs (liver and kidney), fish, shellfish, milk products, eggs, avocados, legumes, mushrooms, and sweet potatoes

• Produced by bacteria in the colon

• Supplements

  • Calcium or sodium pantothenate, or panthenol

  • Multivitamin: 10 mg

  • Single-nutrients: 5-500 mg

Pantothenic Acid Stability

• Stable when dry and in the solution with a neutral pH

• Destroyed

  • heating and freezing

  • acidic and alkaline solutions

• refining of grains, freezing, and canning lowers pantothenic acid content up to 75%

How is Pantothenic Acid found

• present in food in free and bound forms

  • 85% of PA is found as a component of CoA or 4’ phosphopantetheine

Digestion of CoA

• CoA → hydrolyzed by pyrophatase → 4’-phosphopantethine → phosphatase → pantetheine → pantotheinase → pantothenic acid

Pantothenic Acid Absorption

• is absorbed primarily in the jejunum

• ~50% of PA is absorbed (range: 40-61%)

  • High [ ] = Passive diffusion

  • Low [ ] = absorbed by Na-dependent shared multivitamin transporter (SMVT)

  • Panthenol (supplements) - diffusion and converted to PA

• ~10% absorbed with supplement if ingesting 10 x AI (50 mg or more)

Pantothenic Acid Transport, Uptake, and Storage

Transport

• Transported freely in blood from the small intestine enterocyte to liver then the rest of the body

  • Primarily within RBCs

• [PA] = 30 - 60 micrograms/dL

Uptake

• RBC and brain: passive diffusion

• Other tissues: SMVT (in intestine)

Storage

• Present within cells as PA and 4’- phosphospantothenic acid

• Most PA is used to synthesize CoA, which is found in all tissues

  • high [ ] in liver, adrenal gland, kidneys, brain, heart

• Majority of CoA is located in the mitochondria

Pantothenic Acid: Functions and Mechanisms of Actions (CHO)

• Acetic acid → Acetyl CoA, Malonic acid → Malonyl CoA, Propionic acid → Propionyl CoA, Methylmalonic acid → Methylmalonyl CoA, Succinic acid → Succinyl CoA

• CoA is involved in nutrient metabolism: CHO, fat and protein

  • CHO

    • Oxidative decarboxylation of pyruvate to acetyl CoA using pyruvate dehydrogenase complex along with NAD+ to NADH

      • glycolysis

    • Oxidative decarboxylation of A-keto-glutarate to succinyl CoA using using NAD+ to NADH, FAD, thiamin pyrophosphate, and Lipoic acid

      • Kreb’s cycle / TCA cycle

Pantothenic Acid: Functions and Mechanisms of Actions (Lipid Metabolism)

• Synthesis

  • Cholesterol, ketone bodies, fatty acids, phospholipids, steroid hormones, and sphingolipids

• Cholesterol and ketone bodies

  • 2 acetyl-CoA → Acetyl-CoA acetyltransferase → acetoacetyl CoA → HMG-CoA synthase 1 → 3-hydroxy-3-methylglutaryl-CoA → (Rate Limiting Step, needs NADPH as a coenzyme) HMG-CoA reductase → mevalonate → cholesterol

    • statins inhibit the RLS to decrease production of cholesterol

• Keto acid is used to assess ketosis because it has a longer half life

  • glutaryl Co A is important for making ketine bodies

Pantothenic Acid: Functions and Mechanisms of Actions (FA synthesis)

• Condensation of acetyl CoA with activated CO2 to form malonyl-CoA

  • needed for the repetitive addition of 2 carbons on fatty acid synthesis

• Malonyl-CoA binds with ACP and the condensing enzyme combines with acetyl CoA

Pantothenic Acid: Functions and Mechanisms of Actions (Acylation and Acetylation or proteins, sugars, and drugs)

• Acylation and Acetylations

  • Post transitional modification of proteins

    • Acylation: affects protein function, activity, and location

    • Acetylation: affects enzyme activity and function

      • when a acetyl is added to a substrate

  • Acetylation

    • Extensive with proteins - Liver

      • prolongs the ½ life of proteins

    • Usually occurs on lysine residues

    • acetylated aminosugars can provide recognition sites on cell surfaces or direct proteins for membrane functions

    • choline is a acetylated to the neurotransmitter acetylcholine

Pantothenic Acid: Functions and Mechanisms of Actions (Folate)

• 10-formyl tetrahydrofolate dehydrogenase (#10)

  • Requires 4’-phosphopantethiene for activity

  • NADP is the oxidizing agent and CoA is needed

Patonthenic Acid Excretion and Adequate Intake (AI)

• excreted as pantothenate or pantothenic acid

• AI

  • Adults: 5 mg/day

  • Pregnancy 6 mg/day

  • Lactation 7 mg/day

Patothenic Acid Deficinecy

• Unlikely'; occur multiple deficiencies

• Burning Foot Syndrome (rare)

  • numbness of toes and burning sensation in feet and nerve inflammation

  • Exacerbated by warmth and diminished with cold

  • Vomiting, fatigue, muscle weakness, restlessness, irritability

  • Treated with calcium or sodium, pantothenic

• At risk of deficiency

  • alcoholism (low intake/increased excretion), diabetes (increased excretion) inflammatory bowel disease ( decreased absorption)

Pantothenic Acid Toxicity

• No UL

• No toxic level

  • 10 g daily for up to 6 weeks - no side effects

  • > 15 to 20 g associated with mild intestinal distress, including diarrhea

• Assessment of nutriture

  • blood concentration <100 mg/dL may reflect low dietary intake

    • do not correlate well with changes in OA intake and status

    • Urinary pantothenate excretion better indicator of status

      • <1mg/day indications poor PA staus

Biotin Sources

Commonly found free or bound to proteins in liver, soybeans, egg yolk, legumes and nuts

• there is a glycoprotein in raw egg called avidin that binds biotin. It’s an irreversible bond, unless it is cooked

Biotin Digestion

Biotin is bound to protein in food, must be removed from the protein bond

• pepsin in your stomach and proteases either on the brush border or from your pancreas will break the bonds, and you end up with free biotin or biocytin (biotin bound to lysine).

• Biocytin can be further digested to free biotin in the presence of the enzyme biotinidase to biotin and lysine

• there is a genetic mutation that leads to biotinidase deficiency

  • symptoms would be lethargy, hypotonia, seizures, ataxia, dermatitis, and alopecia

Biotin Absorption

• absorbed primarily as free in the proximal small intestine

  • duodenum is the preferred area of absorption

  • In physiological intakes biotin will cross the brush border of the small intestine or the colonic cell membrane with a carrier

    • carrier is a sodium dependent transporter (SMVT) the same transporter for pantothenic acid

    • High levels of biotin suppresses the transcription of the transporter gene

• Undigested biocytin may be absorbed by peptide carriers

  • alcohol decrease absorption

  • biotin synthesized by colonic bacteria is absorbed in proximal and transverse colon (SMVT)

  • transport across the basolateral membrane is carrier-mediated

  • 100% of oral free biotin is absorbed

Biotin Transport, Uptake, and Storage

Transport

• In plasma 80% is free biotin, 20% is bound to proteins (albumin, globulins, and biotinidase)

• Blood (biotin): 200 759 pg/mL

Uptake

• Liver and probably other tissues: SMVT and monocarboxylate transporter (MCT) I

Storage

• Small quantities stored in muscle, liver, and brain

Biotin AI (Adequate Intake) and UL

Adults: 30 micrograms/day

Pregnancy: 30

Lactation: 35

UL: none

• up to 200 mg without side effects

At risk for biotin deficiency

• Those consuming excess raw egg whites

• Those with GI disorders

• People who consume excess alcohol

• Pregnant and lactating women

• Those on anticonvulsant drug

Biotin Assesment of Nutriture

• Decreased urinary biotin excretion (< 6 micrograms/day) is a sensitive and early indicator of biotion deficiency

• a diet devoid of biotin can decrease biotin in plasma and urine in ~2-4 weeks

Biotin as a coenzyme in a holocarboxylase synthetase reaction

• biotin is a covalently attached as a coenzyme to form several holoenzymes

• holoenzymes function as carboxylases

• the carboxylases facilitate the incorporations of a carboxyl group into a substrate

Each holocarboxylase is formed in a reaction catalyzed in two sequential steps by holocarboxylase synthetase

1. formation of biotinyl-AMP from biotin and ATP

2. Formation of an amide bond between the carboxyl group of biotin and the e-amino group on a specific lysine residue in each apocarboxylase with the release of AMP

Biotin and Pyruvate Carboxylase

Role: Converts pyruvate t oxaloacetate

Significance: Replenishes oxaloacetate for TCA cycle. Necessary for glucogenesis. Important for its regulatory function

• Decrease in levels of ATP promotes TCA cycle

• An increase in levels promotes glycogenesis

Biotin and Acetyl-Coa Carboxylase

Role: Forms malonyl-CoA from acetate

Signifigance: Commits acetate units to fatty acid synthesis to start the synthesis

• Rxn needs ATP and biotin

• Acetyl-Coa Carboxylase is considered and regulatory and rate limiting enzyme

• Malonyl CoA will hook up with a acyl carrier protein and acetyl CoA will hook up with a condensing enzyme to create the complex that is important for fatty acid synthesis

Biotin and Propionyl-CoA Carboxylase

Role: Converts propionyl-CoA to methylmalonyl-CoA

Significance: Provides mechanism for metabolism of some amino acids and odd-chain fatty acids to become Propionyl CoA

• Important for converting propionyl CoA into Methylmalonyl CoA

• the catabolism of Threonine, Methionine, Isoleucine, and Valine lead to the product of propionyl CoA

• Beta oxidation of fatty acids involves pulling off 2 C at a time. So when when there is an even # of C the end product is Acetyl CoA

• In order to go to Methylmalonyl CoA requires an enzyme, biotin, magnesium, and ATP as a cofactor

• Methylmalonyl is then converted to Succinyl CoA to be used in the Kreb’s cycle by mutase and B12

Biotin and B-methylcrotonyl-CoA Carboxylase

Role: Converts B-methylcrotonyl-CoA to methylglutaconyl-CoA

Significance: Allows catabolism of leucine and certain isoprenoid compounds

• during catabolism of leucine B-methylcrotonyl-CoA carboxylase

• B-Methylcrotonyl-CoA is carboxylated in the presence of ATP, Mg as a cofactor, and enzyme that is biotin dependent

• There can be a deficiency and the beta methylpropyl choice carboxylase or there could be a decline in activity.

• Therefore this beta methyl crotonyl COA is shunted to other pathways and there are compounds that are made that you could measure in urine that would tell you that this carboxylation reaction is not occurring.

Vitamin B6

• exist as 6 vitamers Pyridoxine (PN; Alcohol form), Pyridoxal (PL; aldehyde form), Pyridoxamine (PM; amine form), Pyridoxine Phosphate (PNP), Pyridoxal Phosphate (PLP) Pyridoxamine Phosphate (PMP)

• PMP is the most active enzyme

Vitamin B6 Sources

Plant Foods

• May be present as a glucoside

• Vegetables (potatoes), some fruits (bananas), and nuts as well fortified cereals

Animal products: PL, PLP, PM, PMP

• Beef, fish, pork, and chicken

Fortified foods and supplements: pyridoxine hydrochloride

• Multi: 2 mg

• single 2-300 mg

Fairly stable with cooking

• Lost in prolonged heat (sterilizing and canning)

• lost in refining/milling of grains (not added back)

• lost in storage

Vitamin B6 Digestion

• PLP, PNP, PMP must be dephosphorylated to PL, PN, PM

• Alkaline phosphatase, a zinc dependent enzyme at brush border and other intestinal phosphatases remove phosphates from PLP, PNP, PMP to make the free forms

Vitamin B6 Absorption

• PL, PN, and PM (the free forms) are absorbed in jejunum by passive diffusion

  • can be absorbed anywhere in GI tract, just prefers the jejunum

  • glucosidases can dephosphorylate the vitamers to the free forms

• 75% (61-97%range) absorbed

• little metabolism in intestinal cells

  • Sometimes pyridoxine can be converted to pyridoxine phosphate or to pyridoxal phosphate

• PN, PL, and PM are released into portal blood and then transported to the liver

• The liver mainly absorbs and metabolizes B6

Vitamin B6 Transport

• PLP and PL are major forms in systemic blood (75-90%)

• Most PLP (and other vitamers) transported bound to albumin

• Liver takes up newly absorbed B6 by passive diffusion

• Oxidase is FMN dependent

• Steps

  • PL and PLP are released from the liver to other tissues

  • PLP is hydrolyzed to PL in blood for cellular uptake

  • PL is then phosphorylated by an intracellular kinase

    • free forms are taken by tissues

  • PLP then binds a protein to prevent its degeneration

    • functions as a coenzyme

Vitamin B6 Storage

• 5-10% in liver

• 75-80% in muscles

  • As PLP bound tp glycogen phosphorylase

    • phosphorylation of B6 traps it in the cell

    • Binding to protein prevents hydrolysis by phosphatases

      • important for adequate amount of B6 to be available

• Other tissues with B6: brain, kidney, spleen

  • typically, phosphorylated and bound to enzymes or PLP binding proteins in the cytosol and mitochondria

Vitamin B6 and Non Coenzyme role (Non PLP role)

Gene expression

• Modulates steroid hormone binding and transcription factor binding to regulatory DNA regions

Vitamin B6 and Coenzymes (primarily as PLP)

• Transamination

• Deamination

• Decarboxylation

• Transulfhydration

• Heme synthesis

• Tryptophan → niacin

• Glycogen degradation

• Transelenation

• Folate

PLP as a coenzyme and reactions will attached by a shift based linkage to the amino group on the enzyme’s active site.

• Is usually lysine

• A shift base is a compound where there’s the presence of a double bond linking C and N

B6 and Transamination

One of the main roles of vitamin B6 is when you have a transfer of an amino group from one amino acid to an alpha ketoacid.

• This makes a new alpha ketoacid and a new amino acid

• The enzyme that is necessary for transamination is called transaminase or Aminotransferase

• PLP is the coenzyme that's important in the involvement of the transferring of the amino group

• EX.

  • Glutamate (keto acid) + pyruvate → a-ketogluterate (keto acid) and alanine (amino acid)

  • Enzyme-PLP Schiff base + amino acid → PMP-Enzyme + a-keto acid product → a keto acid substrate → PLP enzyme

B6 and Demanimation

Is the removal of the amino group

• Amino acid in the presence of dehydrogenase and coenzyme of PLP turn into a Keto acid + NH3

  • a desulfhydrase is used instead if the amino acid contains sulfur

  • NH3 is either converted to urea or added to glutamate to make glutamate or glutamin

    • this is how toxic NH3 is removed

B6 and Decarbxylation

• The elimination of the CO2, the carboxy from a compound usually results in a drop in energy free energy.

Dopamine, epinephrine, norepinephrine

• Tyrosine → Tyrosinase and Biopterin/Vit C → Dihydroxyphenylalanine (DOPA) → Decarboxylase and PLP (Rate Limiting Step)→ Dopamine → Dopamine hydroxylase and Vit C → Norepinephrine → Transmethylase and S-adenosylmethionine → Epinephrine

GABA - is a major inhibitory neurotransmitter, and his primary role is to reduce neuron excitability.

• Glutamate → glutamate decarboxylase and PLP → Y-aminobutyric acid

Serotonin

• Tryptophan → 5-hydroxytryptophan → 5-HTP decarboxylase/aromatic amino acid decarboxylase/PLP → serotonin/5-hydroxytryptamine (5-HT)

Histamine - is a major inhibitory neurotransmitter, and his primary role is to reduce neuron excitability. Ex. important for gastric acid secretion and it regulates vasodilation secretion and bronchoconstriction

• Histidine → Histidine decarboxylase/PLP → histamine

B6 and Transulfhydration

• PLP is required for the enzymes Cystathione B-synthase and Cystathionine y-lase

  • is needed for transulfhydration pathway of methionine to Cysteine

  • methionine is an essential amino acid and when thiamine levels are low cysteine levels become important

B6 and Heme Synthesis

• Glycine + Succinyl Coa → ALA synthase and PLP (Rate Limiting Step) → aminolevulinic acid + CO2 +CoASH

  • occurs in mitochondria

  • zinc and iron are involved

• If there is not an adequate supply of vitamin B6 then there will be impairment in making Heme

  • therefore B6 is associated with some anemias

Vitamin B6 and Tryotophan to Niacin Pathway

• Kynureninase needs B6 (PLP) as a coenzyme)

• Myosin, riboflavin, ATP, and niacin are involved in this process

• If there is not an adequate supply of PLP there will be a build up of 3-OH Kynurenine

B6 and Glycogen Degradation

• PLP is a coenzyme for Glycogen Phosphorylase

  • If there were inadequate amounts of PLP there would be build up of glycogen

• In this process, the phosphate of PLP is thought to stabilize the compound and permit covalent bonding of the phosphate to form glucose 1 phosphate.

B6 and Transelenation

seleneomethionine → Selenocysteine → PLP/B-lyase → Selenide

• Selenide is important for the making of various selenoproteins.

B6 and Folate

• Selene hydroxymethyltransferase requires PLP as a coenzyme to make Tetrahydrofolate (THF) from 5,10-methlyene THF

B6 Metabolism and Excretion

• Vitamin B6 is excreted primarily in urine and very little is excreted in feces

  • 4-pyridoxic acid is major urinary metabolite and is derived from the oxidation of PL

  • The oxidated PL comes from a dehydrogenase that is NAD dependent or an oxidase that is FAD dependent

Vitamin B6 Assessment of Nutriture

• Plasma PLP concentration best indicator of tissue store (best way to asses status)

  • < 5 micrograms/L indicates a vitamin deficiency

  • 5-7.5 indicate marginal status

  • >7.5 indicate adequacy

• Urinary B6 and 4-pyridoxic acid

  • 4-pyridoxic acid: short term indicator of b6 status

• Erythrocyte transaminase activity

Vitamin B6: RDA

• Adults aged 19-50: 1.3 mg/day

• Preganacy: 1.9 mg/day

• Lactation: 2.0 mg/day

• RDA increases with age

  • males aged >51: 1.7 mg/day

  • females aged > 1.5 mg/day

Vitamin B6 Deficiency and Treatment

• Rare in the US

• Deficiency may occur in2-3 weeks, but it may take up to 2.5 months

• Symptoms

  • weakness, fatigue, cheilosis, glossitis, angular stomatitis

  • neurological problems: depression, confusion, peripheral neuropathy, seizures

• Deficiency causes

  • Hypochromic microcytic anemia

  • Impaired Niacin synthesis from tryptophan INhibits Homocysteine metabolism

Treatment

• oral supplements 2.5-25 mg (up to 100 mg) daily for a few weeks

At Risk For B6 Deficiency

• Elderly

  • poor intake, accelerated hydrolysis of PLP, and oxidation of paradoxal

• Excessive alcohol consumption

  • can impair the conversion of paradoxes and pyridoxamine to PLP and the accumulation Of acetaldehyde, which is formed from alcohol metabolism, remember alcohol and the presence of alcohol dehydrogenase and NAD makes acetaldehyde and acetaldehyde. Thus enhances coenzyme degradation

• Systemic inflammation

  • may increase catabolism, and there also may be an increased need for people during inflammation and then malabsorptive conditions.

• Malabsorption condition (inhibits)

  • Medication/Drugs

  • INH (isoniazid)

  • Penicillnamine

  • Corticosteroids - prednisone

  • Anticonvulsants

  • Oral contraceptives

• Vitamin B6 Dependency Syndrome

  • pyridoxine-dependent seizure (PDS)

    • autosomal recessive neurometabolic disorder

Vitamin B6 toxicity

• >200 mg/day: sensory and perpheral neuropathy

  • unsteady gait, paresthesia (tingling/numbness) in extemities, impaired tendon reflexes

• >2 g/day

  • May cause degeneration of neurons in spinal cord and impaired coordination

• UL = 100 mg/day

  • to reduce the risk of developing neuropathy

Vitamin B12 (Cobalamin)

• Macrocylic (corrin) ring made of four reduced pyrrole rings linked together

• Contains cobalt in ring center attached to the nucleotide 5,6 dimethylbenzimidazole

Vitamin B12 Sources

• Primarily from animal products

  • meat and meats products

  • fish and shellfish

  • dairy contains less, nut may be more bioavailable

• Fortified plant based foods (cereals, soymilk, nutritional yeast)

• Fairly stable and resistant to light, heat, oxidation

• Cynocobalamin and hydroxyocobalamin are th main forms in supplements

  • readily cinverted to methylcobalamin and adenosylcobalamin

Vitamin B12 Dijestion

The body absorbs vitamin B12 from food in a two-step process. First, hydrochloric acid and pepsin in the stomach separates vitamin B12 from the protein that it's attached to. Second, the freed vitamin B12 then combines with a protein made by the stomach, called intrinsic factor, in the duodenum, and the body absorbs them together.

Vitamin B12 Absorption

• Carrier mediated intestinal absorption is saturated with 1.5-2 micrograms per meal

• With pharmacological doses, 1-3% absorbed vias passive diffusion through the small intestine (does not need intrinsic factor complex)

• Overall, 50% (11-65% range_ absorbed with usual uptake

  • Absorption efficiency decreases as intake increases

    • protective mechanism so too much is not absorbed

Vitamin B12 Deficiency

• Can take up to 5 years for deficiency to occur

• Enterohepatic Circulation

  • The vitamin is excreted in the bile: however, it can bind to intrinsic factor (IF) in the duodenum and be absorbed in the ileum

    • Related to malabsorption syndrome

Vitamin B12 Transport

• Appears in the blood 3-4 h after absorption and peaks in 8-12 h

  • 60%-80% methylcobalamin and 20% adenosylcobalamin (2 enzyme forms)

• Circulates in blood bound to transcobalamin (TCII) and haptocorrin

  • TCII transports newly absorbed B12 and accounts for 20% of cobalamin in the blood

    • Half-Life: <2hrs

  • Haptocorrin is synthesized in WBCs and transports and accounts 80% of cobalamin in the blood (does not transport newly absorbed B12 but stored B12 )

    • This complex is not taken up by tissues

    • Half life: 10 days

    • Circulating storage and delivering cobalamin from peripheral tissues back to the liver

  • Insertion of arginine of proline diminishes the ability of TCII to bind and transport B12 to tissues

Vitamin B12 Uptake

• Taken up by tissues via tanscobalamin recepeter mediated endocytosis

  • Transport proteins that are in the cell and they're called chaperones and those proteins are thought to carry the vitamin within the cell to the various organelles and various compartments within the cell.

• Metabolized to either coenzyme forms methylcobalamin (cofactor in cytoplasm) or adenosylcobalamin (coenzyme in mitochondria)

Vitamin B12 Storage

• 2-3 mg stored in the body

  • 50% in liver

  • 30% in muscle

  • smaller amounts in pituatary gland, bone, kidneys, heart, brain, spleen

• Mainly as adenosylcobalamin with small amounts if hydroxocobalamin ad methylcobalamin

B12 to make Methionine

• Methionine synthase needs B12 as a coenzyme to make methionine from homocysteine and Cobalamin from Methylcobalamin

  • Methionine synthase takes off the methyl group that methylcobalamin gained from 5-methyl THF and gives it to homocysteine to make methionine

• Cobalamin is easily oxidized meaning methionine synthase easily becomes inactive

  • There is a NADPH-dependent enzyme that reduces methionine synthase to its active form

• Folate is needed to turn THF to 5,10-methylene THF to % methyl THF

  • 5-methyl THF is needed to turn cobalamin into methylcobalamin

B12 and the Oxidation of L- methylmalonyl-CoA

• Startes with a odd #chain fatty acid

• Propionyl → L-methylmalonyl-CoA → Methylmalonic acid or Succinyl CoA

  • Methylmalonic CoA is made without B12

    • Is measured to see if B12 levels are low

  • Succinyl CoA is made with Methylmalonyl CoA mutase which consists units that need 5’deoxyadenosyl cobalamin form of vitamin B12

    • Then enters the Kreb’s cycle

Vitamin B12 Metabolism and Excretion

• Undergoes little to no degradation prior to excretion

• 0.1% is excreted in bile, but (75%) reabsorbed in the ileum

• Only small amounts (~0.25 micrograms/day) lost in urine

• Trace losses through the skin may also occur

Vitamin B12 RDA

• Adults: 2.4 micrograms/day

• Pregnancy: 2.6

• Lactation: 2.8

• Aged>51 fortified or B12 supplements (25-100)

  • due to decrease in consumption of animal products and the ingesting of PPI or antiacids that affects absorption

Vitamin B12 Deficiency

• Megaloblastic Macrocytic Anemia (MA)

• It can be B12 and/or folate deficiency, levels would have to be tested to determine

  • Appears in stages

    • decrease in B12 serum levels

    • decrease in B12 Cellular levels

      • decrease in B12 dependent enzymes

      • decrease in DNA synthesis

      • increase in plasma (homocysteine)

      • increase plasma methylmalonic acid

    • B12 serum levels and plasma methylmalonic acid levels should be measured together

    • Morphological and functional changes occur in RBCs

    • Neurological impairments may also occur

• Pernicious anemia (is only related to B12)

  • autoimmune destruction of gastric parietal and mucosal cells

  • decreased HCl and IF production

B12 AT Risk for Deficiency

• Inadequate intake (vegan; especially infants/children)

• Altered gastric pH

  • Destruction of gastric parietal cells

    • Atrophic gastritis

      • caused by advancing age or by H.pylori infection of stomach

      • loss and inflammation of gastric parietal cells

      • decreased HCl and IF production

• Altered intestinal pH

  • pancreatic impaired exocrine function and Zollinger-Ellison syndrome

  • more acidic pH of small intestine impairs release of B12 from R protein

• Defects in cubam receptors

• Impaired intestinal integrity or function

• Malabsorptive syndromes resections of portions stomach and small intestine

• Competition

  • parasitic infections (tapeworms)

  • Bacterial overgrowth associated with GERD and ulcer medications

Vitamin B12 Deficiency Treatment

• Inadequate intake with neurological symptoms

  • Uptake to 1 mg B12 for 5 weeks followed by lower doses for 1-2 months

• Pernicious anemia and deficiency secondary to malabsorption

  • Monthly IM injections of 500 -1,000 micrograms ot

  • Oral ingestion 1,000-2,000

  • or nasal spray 500

• Improvements may be evident within 10 days

• Serum [metholamlonic] decreases in the first week

• Hematological response: may take up to 2 months

Vitamin B12 Toxicity

• None observed

• No UL

  • excessive amounts can cause GI distress

Vitamin B12 Assessment of Nutriture

• Total serum B12 (B12 TCII + B12 haptocorrin)

  • most common way to measure B12 status

  • you would see low B12 serum and high serum methylmalonic acid and an increase in methyl malonic acid in urine

• Increased serum or urinary methylmalonic acid indicates deficiency

• Increased serum homocysteine> 20 micromol/L if folate and B6 are adequate

• Breath test: subnormal production of labeled CO2 with ingestion of labeled propionate

• Schilling test

  • orally administer radioactive B12 and measure urinary excretion

  • Below normal urinary excretion indicates impaired absorption

• 2 factor when measuring B12

  • when measuring B12 and methylmalonic acid folate should be measured too

  • calcium status is important too bc its responsible for the bonding of intrinsic factor to receptors

Folate

• Folic Acid: Oxidzed form found in fortified foods and supplements

• Folate: Reduced form found in naturally in foods and tissues

Folate Metabolism

Folate Sources

• Mushrooms and green leafy vegetables, brussel sprouts, broccoli, asparagus

• peanuts, beans (especially pento), lentils, stawberries, oranges, cantaloupe

• liver

• Fortified foods: bread, cereal, grain products (140 micrograms/100g)

• Produced by bacteria in colon

Raw foods better than cooked

• Destroyed by heat, oxidation, UV light

• 50-80% lost with certain food processing/preparation

Bioavailability of folate from foods

• Intestinal pH

• Genetic variability in enzymatic activity

• Presence of inhibitors

Folate Forms

Naturally Occuring in foods:

• 5-methyl tetrahydrofolate (THF)

• 5-formyl THF

• 10-formyl THF

• Over 75% attached to multiplr glutamic acid residues

• Overall absorption is about 50% (10%-90% range)

Supplements

• Folic acid (only one glutamic acid residue)

  • monoglutamate

• 5-formyl THF

• %-methyl THF

• Multi: 400 micrograms

• Single ingredient: 1,000 micrograms

• 100% absorbed (especially on empty stomach

• Consumed with food sources absorption is about 85%

Folate RDA

• Adults: 400 micrograms DRE/day

• Pregnancy: 600

• Lactation: 500

Bioavailability

• 1 mcg DFE = 1 mcg food folate

• 1 mcg DFE = 0.6 mcg folic acid from fortified foods or dietary supplements consumed with foods

• 1mcg DFE = 0.5 mcg folic acid from dietary supplements taken on an empty stomach

Folate Absorption

• occurs in the duodenum and upper jejunum

  • bc its transporter (PCFT) prefers a more acidic environment

• Folate or Folic acid → Monoglutamate → PCFT → Enterocyte

  • In the enterocyte, the monoglutamate is methylated at the glutamic acid residue end

  • MRP3 or 5 transports the molecule to the bloodstream

• Pharmacological doses of folic acid are absorbed through passive diffusion

• In the colon uses the transporter RFC

Folate Digestion

• Polyglutamate →glutamate carboxypeptidase and zinc → monoglutamate

• Enzyme is affected by

  • zinc deficiency

  • Acidic pH

  • Alcohol ingestion

  • inhibitors in legumes, cabbage, oranges

• Folic acid in fortified foods and supplements already in monoglutamate form

Folate Transport

• Blood

  • Folate is bound as free monoglutamate (~1/3) or bound to proteins (~2/3) such as albumin and macroglobulins and folate binding protein

  • found primarily as 5-methyl THF and smaller amounts of THF, 10formyl THF, and other THF derivatives

• Higher [folate] in cerebral spinal fluid and RBCs than plasma itself

• Folate in RBCs attained during erythopoiesis: index of long-term folate status (2-3 months)

  • Plasma levels represent folate intake

Folate Uptake

• uptake of folate into tissues: carrier mediated

  • PCFT: liver, pancreas, kidney, and spleen

  • RFC: folate, especially 5-methyl THF for systemic circulation

  • Organic anion transporting poly pepetides (OATP) B1 and B3: liver

  • Folate receptors mediate uptake via endocytosis in tissues including the brain

    • mutation prevent folate from crossing the BBB

    • low cerebral spinal fluid [folate] and neurological problems

Folate Hepatic Metabolism

THF in hepatocytes

• THF (33%)

• 5-methyl THF (~33%)

• 5- and 10-formyl THF (~33%)

Folic acid → DHF → THF (cytosol)

• NADPH dependent DHF reductase

• THF and THF derivatives: glutamate residues added (one at a time) by folylpolyglutamate synthase (ATP dependent)

• Traps THF in hepatocytes → prevents degradation and enables storage

• y-glutamyl hydrolases remove glutamate residues before release into the blood or secreted into bile

  • 50% secreted into bile by MRP2 and breast cancer resistant protein (BCRP) carriers

Folate Storage

• Body stores: 7-30 mg

  • Half of body’s folate is found in the liver

  • Stored wth intracellular folate-binding proteins

  • Main storage forms:

    • Polyglutamate forms of THF and 5-methyl THF

Folate Functions

• One-carbon metabolism

  • Accepts and donates one carbon units

    • Amino acid metabolism

    • DNA synthesis metabolism

      • Cells with short lifespans are particularly affected by folate inadequacy

    • Methyl group (CH3) carrier

      • Impacts gene expression

• Tetrahydrofolate functions as a coenzyme in the mitochondria and the cytosol

  • THF is generated from amino metabolism

• THF derivates serve as donors of one carbon units in synthetic reactions such as amino acid, purine, and pyrimidine synthesis

• Poor folate statues is associated with decreased DNA methylation

Folate and Disease

• Low folate intake associated with increased plasma [homocysteine]

  • Increase in [homocysteine] associated with premature heart disease and stroke

  • cognitive dysfunction

• Folate deficiency associated with increased initiation of colon ( and other) cancer

  • can also inhibit gene translation

• decrease serum and RBC [folate] associated with depression

• Neural tube defects

  • Spina bifida

  • anencephaly

Folate Metabolism and Excretion

• Excreted in urine intact and as metabolites

• In kidneys, folate binding proteins in renal brush border membrane aids tubular reabsorption

• Folate is secreted by liver into bile

  • most is reabsorbed with enterohepatic recirculation

• Folate losses in feces are minimal

• Bacteria in intestine can synthesize folate, and that can be excreted in feces

Folate Deficiency

• Megaloblastic macrocytic anemia (MA)

  • few, abnormally nucleated (immature) and large RBCs

  • fatigues, weakness, headaches, irritability, difficulty concentrating, shortness of breath, heart palpitations

  • can rise from deficiency of folate or B12

  • disrupt DNA synthesis (replication) and cell division

  • Purine and pyrimidine synthesis compromised → macrocytic and megaloblastic cells

  • Treatment: 1-5 mg (oral ingestion) folate daily

    • 5 methyl THF more effective than folic acid

• Diagnosis and progression

  • 1-2 months: decrease plasma [folate] and increase [homocysteine

  • 3-4 months: decrease in RBC [folate]

  • 4-5 months: rapidly dividing cells become megaloblastic

  • increase MCV, hyper segmentation of WBC and decrease blood cell count

    • blood’s oxygen-carrying capacity

At Risk for Folate Deficiency

• Excessive alcohol ingestion

• Malabsorption disorders

• Gastric bypass

• Medications

  • diuretic

  • anticonvulsants

  • methotrexate

  • cholestyramine

  • sulfasalazine

Folate Toxicity

• UL = 1,000 mcg synthetic (non-natural) folic acid

• Folate can mask B12 deficiency

  • Folic acid supplements can alleviate the MA caused by B12 deficiency, but the neurological damage progresses undetected and is irreversible

• 15 mg folate daily

  • insomnia, malaise, irritability, GI distress

• Some studies show increased risk of cancer with 1,00 mcg folic acid daily

Folate Assessment of Nutriture

• Plasma, serum, or RBC concentration

• Serum or plasma [folate] reflect recent dietary intake

  • True deficiency diagnoses requires repeated measures of serum/plasma folate

• RBC [folate] better indicator of folate status

• Formiminoglutamate excretion (FIGLU) excretion

• A functional marker of folate and vitamin B12 deficiencies is elevated plasma homocysteine concentration