Water solubility conjugate transport

Many of the phase 1 enzymes are located in hydrophobic membrane environments. In vertebrates, they are particularly associated with the endoplasmic reticulum of the liver, in keeping with their role in detoxication. Lipophilic xenobiotics are moved to the liver after absorption from the gut, notably in the hepatic portal system of mammals. Once absorbed into hepatocytes, they will diffuse, or be transported, to the hydrophobic endoplasmic reticulum. Within the endoplasmic reticulum, enzymes convert them to more polar metabolites, which tend to diffuse out of the membrane and into the cytosol. Either in the membrane, or more extensively in the cytosol, conjugases convert them into water-soluble conjugates that are ready for excretion. Phase 1 enzymes are located mainly in the endoplasmic reticulum, and phase 2 enzymes mainly in the cytosol. 

This transporter is particularly important in the small intestine, in the gut wall enterocytes, where its activity in humans is sevenfold higher than liver tissue. In the gut, pGp, acting in concert with cytochrome P-450 (CYP3A4) (see chap. 4), functions to keep chemicals, which may be potential toxicants, out of the body by pumping them back into the lumen of the gut. The CYP3A4 converts them into more polar compounds, which are less readily absorbed or further metabolized into water-soluble conjugates. 

The majority of xenobiotics that enter the body tissues are lipophilic, a property that enables them to penetrate lipid membranes and to be transported by lipoproteins in body fluids. The metabolism of xenobiotics, carried out by a number of relatively nonspecific enzymes, usually consists of two phases. During phase I, a polar group is introduced into the molecule and although this increases the molecule s water solubility, the most important effect is to render the xenobiotic a suitable substrate for phase II reactions. In phase II reactions, the altered compounds combine with an endogenous substrate to produce a water-soluble conjugation product that is readily excreted. Although this sequence of events is generally a detoxication mechanism, in some cases the intermediates or final products are more toxic than the parent compound, and the sequence is termed an activation or intoxication mechanism. See Chapter 20 for discussion of activation and toxicity. 

Bilirubin is produced by the transformation of haem (mainly from the destruction of red blood cells) via biliverdin (see Figure 2.8). This takes place in the liver, spleen and bone marrow. Bilirubin is transported to the liver in the serum attached to albumin, and at this stage is unconjugated. It is insoluble in water and hence cannot be excreted in this form. Hepatocytes transform unconjugated bilirubin into a water-soluble conjugated form which is excreted via the bile into the intestine. Here, some is converted to urobilinogen and excreted by the kidneys, the majority being converted to stercobilin and excreted in the faeces. 

The effect of temperature on fluorescence has been studied, as has the effect of salt concentration and water-soluble conjugated polymers. A method for the quantification of ssDNA dsDNA is described, as well as kinetics of mismatch hybridization and the kinetics of collision in short ss-nucleic acids. Fluorescence quenching of Cy-5 labelled oligonucleotides by poly(phenylene ethynylene) particles has been shown to be a more sensitive method than excitation of the Cy-5 fluorophore. An ultrasensitive method for the detection of DNA uses highly fluorescent conjugated nanoparticles, and detection limits below IfM were achieved. DNA transport through a carbon nanotube has also been observed using fluorescence microscopy. " ... 

The water soluble conjugate is finally eliminated from the cell by transport proteins (organic anion transporters, multidrug resistance associated proteins), and finally excreted via the renal or the bile route. This transport step is considered as phase HI of drug metabolism. It does not require further biotransformation, and will be not considered in this chapter. 

The bulk of the bilirubin produced in the human organism arises in cells of the reticuloendothelial system by the metabolic degradation of hemoglobin. The pigment is released into the blood, and is transported to the liver as a bilirubin-albumin complex. Enzymic processes in the liver convert the very hydrophobic bilirubin into water-soluble conjugates. This is a key step in the excretion of bilirubin, since it allows for the effective concentration of this compound in the bile. 

Glucuronide conjugates are ionized at urinary pH and being water soluble can be readily eliminated. They are also substrates for the organic anion transporter. 

Polymeric micelles formed by Pluronics, PEG phospholipid conjugates, PEG-b-polyesters, or PEG-b-poly-L-amino acids were proposed for drug delivery of poorly water-soluble compounds, such as amphotericin B, propofol, paclitaxel, and photosensitizers [77,86,87]. It was also emphasized that using polymeric micelles can significantly increase the drug transport into the brain. 

Figure 22-3. Transport and hepatic metabolism of bilirubin. Bilirubin that is produced in phagocytes is transported to liver as an albumin-bilirubin complex. Uptake into the hepatocytes takes place in liver sinusoids. Within the hepatocyte, bilirubin is transported to the endoplasmic reticulum (microsomes) bound to glutathione S-transferase (GST). Bilirubin is made water soluble by addition of one or two glucuronic acid moieties obtained from UPD-glucuronic acid, catalyzed by bilirubin-UDP-glucuronyltransferase. The product, conjugated bilirubin, is transported across the bile canalicular membrane for secretion into the biliary system, with subsequent movement into the intestines.

Conjugation of lipophilic xenobiotics to polar cellular constituents renders the xenobiotic more water-soluble. While the lipophilic parent xenobiotics could readily diffuse into the cells, the increase in polarity associated with conjugation greatly reduces the ability of the compound to diffuse across the lipid bilayer of the cell membrane thus trapping the compound within the cell. The polar conjugates must therefore rely upon active transport processes to facilitate efflux from the cell. Hepatocytes, as well as other cells involved in chemical detoxification, are rich with members of the ATP-binding cassette superfamily of active transport proteins (ABC transporters). Cellular efflux of xenobiotics by these transporters is often referred to as Phase III elimination because Phase I or II detoxification processes often precede and are a requirement of Phase III elimination. A detailed description and discussion of elimination and transporters is presented in Chapter 15. 

Modulation of liver and kidney function. Nutrients and xenobiotics (such as secondary metabolites) are transported to the liver after resorption in the intestine. In the liver, the metabolism of carbohydrates, amino acids, and lipids takes place with the subsequent synthesis of proteins and glycogen. The liver is also the main site for detoxification of xenobiotics. Lipophilic compounds, which are easily resorbed from the diet, are often hydroxylated and then conjugated with a polar, hydrophilic molecule, such as glucuronic acid, sulfate, or amino acids (312). These conjugates, which are more water soluble, are exported via the blood to the kidney, where they are transported into the urine for elimination. 

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