Vitamin mobilization

Dihydroxyvitamin (283) is the endogenous ligand for the vitamin receptor (VDR). It modulates genomic function in a tissue and developmentaHy specific manner and affects ceU proliferation, differentiation, and mineral homeostasis (74). Vitamin mobilizes calcium from the bone to maintain plasma Ca " levels. Vitamin and VDR are present in the CNS where they may play a role in regulating Ca " homeostasis. Vitamin D has potent immunomodulatory activity in vivo. 

Vitamins A, D, and E are required by mminants and, therefore, their supplementation is sometimes necessary. Vitamin A [68-26-8] is important in maintaining proper vision, maintenance and growth of squamous epitheHal ceUs, and bone growth (23). Vitamin D [1406-16-2] is most important for maintaining proper calcium absorption from the small intestine. It also aids in mobilizing calcium from bones and in optimizing absorption of phosphoms from the small intestine (23). Supplementation of vitamins A and D at their minimum daily requirement is recommended because feedstuffs are highly variable in their content of these vitamins. 

Mobilization and Metabolism. The total ascorbic acid body pool in healthy adults has been estimated to be approximately 1.5 g, which increases to 2.3—2.8 g with intakes of 200 mg/d (151—158). Depletion of the body pool to 600 mg initiates physiological changes, and signs of clinical scurvy are reported when the body pool falls below 300 mg (149). Approximately 3—4% of the body pool turns over daily, representing 40—60 mg/d of metabolized, or consumed, vitamin C. Smokers have a higher metaboHc turnover rate of vitamin C (approximately 100 mg/d) and a lower body pool than nonsmokers, unless compensated through increased daily intakes of vitamin C (159). The metaboHsm of ascorbic acid varies among different species. 

Metabolism and Mobilization. On entry of vitamin B 2 into the cell, considerable metaboHsm of the vitamin takes place. Co(III)cobalamin is reduced to Co(I)cobalamin, which is either methylated to form methylcobalamin or converted to adenosylcobalamin (coenzyme B>22)- The methylation requires methyl tetrahydrofolate. 

Hydroxy vitamin D pools ia the blood and is transported on DBF to the kidney, where further hydroxylation takes place at C-1 or C-24 ia response to calcium levels. l-Hydroxylation occurs primarily ia the kidney mitochondria and is cataly2ed by a mixed-function monooxygenase with a specific cytochrome P-450 (52,179,180). 1 a- and 24-Hydroxylation of 25-hydroxycholecalciferol has also been shown to take place ia the placenta of pregnant mammals and ia bone cells, as well as ia the epidermis. Low phosphate levels also stimulate 1,25-dihydtoxycholecalciferol production, which ia turn stimulates intestinal calcium as well as phosphoms absorption. It also mobilizes these minerals from bone and decreases their kidney excretion. Together with PTH, calcitriol also stimulates renal reabsorption of the calcium and phosphoms by the proximal tubules (51,141,181—183). 

Although it is being found that vitamin D metaboUtes play a role ia many different biological functions, metaboHsm primarily occurs to maintain the calcium homeostasis of the body. When calcium semm levels fall below the normal range, 1 a,25-dihydroxy-vitainin is made when calcium levels are at or above this level, 24,25-dihydroxycholecalciferol is made, and 1 a-hydroxylase activity is discontiaued. The calcium homeostasis mechanism iavolves a hypocalcemic stimulus, which iaduces the secretion of parathyroid hormone. This causes phosphate diuresis ia the kidney, which stimulates the 1 a-hydroxylase activity and causes the hydroxylation of 25-hydroxy-vitamin D to 1 a,25-dihydroxycholecalciferol. Parathyroid hormone and 1,25-dihydroxycholecalciferol act at the bone site cooperatively to stimulate calcium mobilization from the bone (see Hormones). Calcium blood levels are also iafluenced by the effects of the metaboUte on intestinal absorption and renal resorption. 

The following polyvitamin prepai ations were analyzed Kal tsid (OAO Comfort Plus , Russia), Asvitol (OAO INC Marbiofarm , Russia), Pikovit (KRKA, d.d. The New Place, Slovenia), Yeast with vitamin C (000 EKKO Plus , Russia). Chromatographic experiment has been carried out using Silufol UV-254 (Kavalier, Czech Republic) and acetone - ethyl acetate - acetic acid - ethanol (3 5 1 1) - CTAB (2T0 M) as a mobile phase mixture. The linearity calibration plot, built in coordinate S = f (IgqAC), is valid in the interval 5-25 p.g. Correctness of the determination has been checked by photometry. The obtained results for the ascorbic acid determination are presented below. 

Other microporous materials have been synthesized using the porogen polyethylene glycol in polyethylene oxide-urethane gels [27]. Micropores were formed in the gel, and it was found that the diffusion of larger species, vitamin B12, was enhanced relatively more than that of a smaller species, proxyphylline. This result is in qualitative agreement with that found for electrophoretic transport by RiU et al. [322] discussed earher, where the mobility of larger species was preferentially enhanced in the templated media. 

Ubiquinone or Q (coenjyme Q) (Figure 12-5) finks the flavoproteins to cytochrome h, the member of the cytochrome chain of lowest redox potential. Q exists in the oxidized quinone or reduced quinol form under aerobic or anaerobic conditions, respectively. The structure of Q is very similar to that of vitamin K and vitamin E (Chapter 45) and of plastoquinone, found in chloroplasts. Q acts as a mobile component of the respiratory chain that collects reducing equivalents from the more fixed flavoprotein complexes and passes them on to the cytochromes. 

Reverse phase chromatography is finding increasing use in modern LC. For example, steroids (42) and fat soluble vitamins (43) are appropriately separated by this mode. Reverse phase with a chemically bonded stationary phase is popular because mobile phase conditions can be quickly found which produce reasonable retention. (In reverse phase LC the mobile phase is typically a water-organic solvent mixture.) Rapid solvent changeover also allows easy operation in gradient elution. Many examples of reverse phase separations can be found in the literature of the various instrument companies. 

Anemia may be present in some patients due to impaired erythropoietin regulation, nutritional factors (vitamin E and iron malabsorption), or chronic inflammation. With chronic pulmonary disease, increased cytokine production can lead to shortened red blood cell survival, reduced erythropoietin response, and impaired mobilization of iron stores. 

Stimulating activation of vitamin D by 1-a-hydroxylase to cal-citriol (1,25-dihydroxyvitmin D3) to promote calcium absorption in the GI tract and increased calcium mobilization from bone... 

Factors that can predispose patients to developing metabolic bone disease include deficiencies of phosphorus, calcium, and vitamin D vitamin D and/or aluminum toxicity amino acids and hypertonic dextrose infusions chronic metabolic acidosis corticosteroid therapy and lack of mobility.35,39 Calcium deficiency (due to decreased intake or increased urinary excretion) is one of the major causes of metabolic bone disease in patients receiving PN. Provide adequate calcium and phosphate with PN to improve bone mineralization and help to prevent metabolic bone disease. Administration of amino acids and chronic metabolic acidosis also appear to play an important role. Provide adequate amounts of acetate in PN admixtures to maintain acid-base balance. 

The a ns wer is a. (Hardman, pp 1525-1528.) Pa r a thyroid ho r m o ne is synthesized by and released from the parathyroid gland increased synthesis of PTI1 is a response to low serum Ca concentrations. Resorption and mobilization of Ca and phosphate from bone are increased in response to elevated PTI1 concentrations. Replacement of body stores of Ca is enhanced by the capacity of PTH to promote increased absorption of Ca by the small intestine in concert with vitamin D, which is the primary factor that enhances intestinal Ca absorption. Parathyroid hormone also causes an increased renal tubular reabsorption of Ca and excretion of phosphate. As a consequence of these effects, the extracellular Ca concentration becomes elevated. 

Karpowich, N. K., Huang, H. H., Smith, P. C. and Hunt, J. F. (2003). Crystal structures of the BtuF periplasmic-binding protein for vitamin B12 suggest a functionally important reduction in protein mobility upon ligand binding, J. Biol. Chem., 278, 8429-8434. 

The overall metabolism of vitamin A in the body is regulated by esterases. Dietary retinyl esters are hydrolyzed enzymatically in the intestinal lumen, and free retinol enters the enterocyte, where it is re-esterified. The resulting esters are then packed into chylomicrons delivered via the lymphatic system to the liver, where they are again hydrolyzed and re-esterified for storage. Prior to mobilization from the liver, the retinyl esters are hydrolyzed, and free retinol is complexed with the retinol-binding protein for secretion from the liver [101]. Different esterases are involved in this sequence. Hydrolysis of dietary retinyl esters in the lumen is catalyzed by pancreatic sterol esterase (steryl-ester acylhydrolase, cholesterol esterase, EC 3.1.1.13) [102], A bile salt independent retinyl-palmitate esterase (EC 3.1.1.21) located in the liver cell plasma hydrolyzes retinyl esters delivered to the liver by chylomicrons. Another neutral retinyl ester hydrolase has been found in the nuclear and cytosolic fractions of liver homogenates. This enzyme is stimulated by bile salts and has properties nearly identical to those observed for... 

Ion-pairing reagents are detergent-like molecules added to the mobile phase to provide additional retention or selectivity for the analytes with opposite charge. Long-chain alkyl sulfonates are commonly used for the separation of water-soluble basic analytes as shown in Figure 16 in the analysis of water-soluble vitamins (WSV). Hexanesulfonate binds with... 

FIGURE 16 HPLC chromatogram of water-soluble vitamins using ion-pair chromatography. LC conditions and peak identification are shown in the inset.The retention times of basic analytes (pyridoxine and thiamine) are strongly dependent of the concentration of ion-pairing reagent (1-hexanesulfonate) in the mobile phase. Reprinted with permission from Reference 17. 

Much discussion of vitamin D focuses on bone health, though this is by no means the only focus on vitamin D action. One result of l,25(OH)2D action is the upregulation of the synthesis of a calcium-binding protein whose function is to transport dietary calcium across the intestinal mucosa and into the systemic circulation. Phosphate accompanies the calcium. This has the effect of increasing the fraction of dietary calcium that is actually absorbed and is, therefore, potentially useful for bone formation. In addition, l,25(OH2)D has the effect of mobilizing calcium from bone. Both actions tend to raise the extracellular level of calcium. 

The role of ubiquinone (coenzyme Q, 4) in transferring reducing equivalents in the respiratory chain is discussed on p. 140. During reduction, the quinone is converted into the hydroquinone (ubiquinol). The isoprenoid side chain of ubiquinone can have various lengths. It holds the molecule in the membrane, where it is freely mobile. Similar coenzymes are also found in photosynthesis (plastoquinone see p. 132). Vitamins E and K (see p. 52) also belong to the quinone/hydroquinone systems. 

The effects of the steroid hormone calcitriol (see p. 330) in bone are complex. On the one hand, it promotes bone formation by stimulating osteoblast differentiation (top). This is particularly important in small children, in whom calcitriol deficiency can lead to mineralization disturbances (rickets see p.364). On the other hand, calcitriol increases blood Ca "" levels through increased Ca "" mobilization from bone. An overdose of vitamin D (chole-calciferol), the precursor of calcitriol, can therefore have unfavorable effects on the skeleton similar to those of vitamin deficiency (hypervitaminosis see p.364). 

Water-soluble vitamins in formulations have been determined by use of ion-pair chromatography. The vitamins include several B vitamins as well as niacin, folic acid, and ascorbic acid (565). Vitamins D and Da were rapidly separated on reverse phase columns (247) as are vitamins A, D, and E in multivitamin tablets (564). Addition of silver ions to the mobile phase has been shown to increase the flexibility inherent in RPC by complexing with the unsaturated bonds and thereby decreasing the retention factor. This effect is also observed with other unsaturated drug molecules including steroids (247). Vitamin A and related compounds have... 

Analysis of vitamin content of food materials appears to be a developing field. B vitamins in rice were analyzed using a mobile phase which contained pentanesulfonic acid and heptanesulfonic acid (558). Although the peaks were not sharp, the separation of the vitamins was satisfactory. Vitamin D in fortified milk has b n analyzed after removal Of cholesterol and carotenes in a preliminary cleanup (559, 540). Vitamin A has been analyzed in margarine, infant formula, and fortified milk (541, 542). Reports of the analysis of other vitamins in food are few to te but this mode of analysis can be expected to rapidly expand in the future in light of the variety of vitamin determinations in formulations which have been done (see Section VIII,F,l). 

Additionally, a further study described an increased catabolism and mobilization of vitamin A in the whole body (Kelley et al, 1998). 

Both NP (428, 441, 442) and RP (427, 436, 437) chromatography has been applied to analysis of vitamin A and provitamin A. A Lichrosorb Si60 column (250 mm x 4.6 mm) has been used for NP chromatography using hexane and hexane/2-propanol as mobile phase [441], More widespread are C8, C18, and C30 column with acetonitrile, methanol, water, and mixtures of these as the most used mobile phase [424,427,428,443], Chavez-Servm et al. [442] reported the use of a short narrow-bore column (50 mm X 2 mm id), which enables less solvent consumption and higher mass sensitivity. 

When fluorescent detection is used, the mobile phase contains zinc chloride as the reduction agent for vitamin K derivatization. The most used mobile phases are methanol and dichloromethane or water. 

RP chromatography is used for analytical determination of vitamin Bg, such as ODS, C18, and amide. The mobile phases used are, like for vitamin Bj and B2, acetonitrile, methanol, and water within a percentage of buffer. 

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