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. 2010 May 27;53(10):4259-65.
doi: 10.1021/jm100254w.

Colloid Formation by Drugs in Simulated Intestinal Fluid

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Free PMC article

Colloid Formation by Drugs in Simulated Intestinal Fluid

Allison K Doak et al. J Med Chem. .
Free PMC article

Abstract

Many organic molecules form colloidal aggregates in aqueous solution at micromolar concentrations. These aggregates promiscuously inhibit soluble proteins and are a major source of false positives in high-throughput screening. Several drugs also form colloidal aggregates, and there has been speculation that this may affect the absorption and distribution of at least one drug in vivo. Here we investigate the ability of drugs to form aggregates in simulated intestinal fluid. Thirty-three Biopharmaceutics Classification System (BCS) class II and class IV drugs, spanning multiple pharmacological activities, were tested for promiscuous aggregation in biochemical buffers. The 22 that behaved as aggregators were then tested for colloid formation in simulated intestinal fluid, a buffer mimicking conditions in the small intestine. Six formed colloids at concentrations equal to or lower than the concentrations reached in the gut, suggesting that aggregation may have an effect on the absorption and distribution of these drugs, and potentially others, in vivo.

Figures

Figure 1
Figure 1
Chemical structures of drugs tested for aggregation in biochemical buffers.
Figure 2
Figure 2
Autocorrelation curves from DLS showing (A) drugs in 50 mM phosphate buffer and (B) in FeSSIF: (●) KPi, (▼) tocopherol nicotinate, (◆) etravirine, (▲) delavirdine, (◼) menatetrenone, (▽) itraconazole, (△) methylene blue. (C) Calibration beads in FeSSIF: (●) FeSSIF, (▼) 15 fM beads, (◆) 60 fM beads, (▲) 240 fM beads, (◼) 480 fM beads. Each curve is one representative sample from each set.
Figure 3
Figure 3
UV−visible quantification of colloids pelleted from FeSSIF by centrifugation. Bars illustrate the percent found in the (◻) supernatant and (◼) pellet: (a) nicardipine (60 μM), a previously described strong aggregator, in 50 mM KPi; (b) nicardipine (60 μM), which is not observed to form colloids in FeSSIF; (c) itraconazole (35 μM); (d) delavirdine mesylate (750 μM).
Figure 4
Figure 4
β-Lactamase cosediments with colloids in FeSSIF. Lane 1 is 2 μg of β-lactamase in 1 mL of FeSSIF, without centrifugation. Lanes 2 and 3 are the supernatant (S) and pellet (P), respectively, from centrifugation of β-lactamase alone. Lanes 4 and 7 are the pellets from centrifugation of each colloid-forming drug alone. Lanes 5 and 8 are the supernatants, and lanes 6 and 9 are the pellets of β-lactamase incubated with colloid forming drugs.
Figure 5
Figure 5
Negative stain electron microscopy of colloid-forming compounds. (A) Phosphate buffer is shown with homogeneous background without particulates. (B) Itraconazole in phosphate buffer forms large colloids, while under the same conditions methylene blue (C) and delavirdine mesylate (D) form small and intermediate sized colloids, respectively. (E) Negatively stained FeSSIF reveals only low-contrast lipid and detergent structures based on the taurocholate and lecithin content. Itraconazole (F), methylene blue (G), and delavirdine mesylate (H) retain their colloid forming ability in FeSSIF, forming colloids similar to those seen in phosphate buffer. The contrast for panels B, C, D, G, and H was adjusted nonlinearly by using a high-pass Fourier filter to reduce the intensity of the negative stain. Bar = 100 nm.

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