In Vitro ABC Transporter Assays for Nutrient Absorption and Drug-Nutrient interaction studies

November, 2005

Table of contents:

1. Digestion 2. Transport of nutrients 3. Importance of ABC transporters in nutrition 4. SOLVO technology 5. References

1. Digestion {1}

Digestion involves the mixing of food, its movement through the digestive tract, and the chemical breakdown of the large molecules of food into smaller ones (Table 1). Digested food molecules, water and minerals are absorbed from the cavity of the upper small intestine and then carried off in the bloodstream to other parts of the body for storage or further chemical change (Figure 1). Nutrients are substances contained in foods which provide energy and raw materials for the synthesis and maintenance of living matter. Human nutrients are proteins, carbohydrates, fat, minerals, vitamins and water. All cells acquire the molecules and ions they need from their surrounding extracellular fluid (ECF). There is an unceasing traffic of molecules and ions: · In and out of the cell through the plasma membrane; · In eukaryotic cells, there is also transport in and out of membrane-bounded intracellular compartments such as the nucleus, endoplasmic reticulum, and mitochondria.

Figure 1: The bloodstream

2. Transport of nutrients{20}

Nutrients can be transported through the cell membrane in 3 ways (Figure 2): Passive diffusion consists of the transport of water and water-soluble substances and small lipids through the lipid bilayer with a concentration gradient. In the case of a facilitated diffusion, transporter proteins create a water-filled pore through which ions and small hydrophilic molecules can pass by diffusion. Fructose, riboflavin and vitamin B12 (in combination with intrinsic factor) are among the substances absorbed by facilitated diffusion. During active transport, transmembrane proteins, called transporters, use the energy of ATP to force ions or small molecules through the membrane against their concentration gradient. These active transport mechanisms have been identified for intestinal absorption of many substances including glucose, galactose, amino acids, calcium, iron, folic acid, ascorbic acid, thiamin and bile acids. Transport proteins, embedded in lipid membranes, facilitate the import of nutrients into cells or the release of toxic products into the surrounding medium. The most important family of membrane transport proteins are the ATP-binding cassette (ABC) transporters. These ABC proteins play a central role in all living cells in the nutrient uptake, protein, drug and antibiotic excretion, osmoregulation, antigen presentation, signal transduction and other important cellular functions.

Figure 2: Passive diffusion, facilitated diffusion and active transport of nutrients

3. Importance of ABC transporters in nutrition {1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19}

Some studies suggest that dietary constituents regulate the expression of ABC Transporters. Changes in ABC Transporter expression may represent an important physiological response to foods containing toxins and an important component of the acute phase immune response. It has been shown that dietary phytochemicals have inhibitory effects on P-glycoprotein (MDR1) and potencies to cause drug-food interactions. Moreover, the observations in a number of studies demonstrated important roles of membrane transporters, i.e. Multidrug Resistant Protein 1 (MRP1), Multidrug Resistant Protein 2 (MRP 2), Breast Cancer Resistance Protein BCRP (MXR) and P-glycoprotein (MDR1) in the cellular accumulation, transport and potential effects of many nutrients. Since some of these nutrients are found in fruits and vegetables, their effect on MRP1, MRP2, MXR and MDR1 may be a mechanism relevant to carcinogenesis and the observed lowered cancer risk in humans with higher dietary intake of fruits and vegetables (Table 1). Table 1: Nutrient-transporter interactions

4. SOLVO Technology

The screening of the interaction of nutrients with ABC transporters is an excellent way to determine the transport of nutrients and also to study drug-food interactions. It is highly recommended to perform screens for MRP1, MRP2, MXR (BCRP) and MDR1. It is also an option to screen for other transporters as well. Detailed references and the description of the assays are available at www.solvo.com. A wide array of ABC transporter assays is available from SOLVO for the different ABC transporters. The patented Calcein Assay, ATPase Assay and some Vesicular transport assays are optimized for high-throughput screening on 96 well plates and widely used by pharmaceutical companies. SOLVO provides full technology transfer for in-house use as well as fee-for-service screening.

5. References

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