Pyranine (8-hydroxy-1,3,6-pyrenetrisulfonate) as a probe of internal aqueous hydrogen ion concentration in phospholipid vesicles

Journal Article · Thu Jan 01 04:00:00 EST 1981 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00509a019· OSTI ID:7090061

Clement, N R; Gould, J M

The fluorescence intensity (at 510 nm) of the hydrophilic pyrene analogue 8-hydroxy-1,3,6-pyrenetrisulfonate (pyranine) is strongly dependent upon the degree of ionization of the 8-hydroxyl group (pK/sub a/ = 7.2) and hence upon the medium pH, over the range pH 6-10. Because of its polyanionic character, pyranine does not bind significantly to phospholipid vesicles having a net anionic surface charge. As a result, it is possible to form vesicles in the presence of pyranine which, after removal of external probe by gel filtration, contain pyranine entrapped within the internal aqueous compartment. Once entrapped, pyranine does not readily leak out of the vesicles. Because the fluorescence properties of entrapped pyranine resemble closely the properties of bulk pyranine solution with respect to pH sensitivity, pyranine can be used as a reliable reporter of aqueous pH changes within anionic vesicles. When HCl is rapidly added to a suspension of unilamellar soybean phospholipid (asolectin) vesicles preincubated at alkaline pH, a biphasic decrease in the pH of the vesicle inner aqueous compartment is observed. An initial, very rapid and electrically uncompensated H/sup +/ influx (t/sub 1/2/ < 1 s) results in the generation of a transmembrane electric potential opposing further H/sup +/ influx. This leads to the development of a much slower (t/sub 1/2/ approx. = 5 min), valinomycin-sensitive, proton-counterion exchange which continues until the proton concentration gradient is eliminated. Similar results were obtained in asolectin vesicles prepared by detergent dilution, in sonicated egg phosphatidylcholine vesicles, and in multilamellar asolectin liposomes.

• Research Organization: Univ. of Notre Dame, IN
• OSTI ID: 7090061
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 20:6; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550200* -- Biochemistry
59 BASIC BIOLOGICAL SCIENCES
AROMATICS
CELL CONSTITUENTS
CELL MEMBRANES
CONDENSED AROMATICS
ESTERS
FLUORESCENCE
HYDROCARBONS
LIPIDS
LIPOSOMES
LUMINESCENCE
MEASURING METHODS
MEMBRANES
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANIC SULFUR COMPOUNDS
ORGANOIDS
PERMEABILITY
PH VALUE
PHOSPHOLIPIDS
PYRENE
SULFONIC ACID ESTERS