Determination of the Acid Dissociation Constant of the Biosurfactant Monorhamnolipid in Aqueous Solution by Potentiometric and Spectroscopic Methods

Abstract

The acid dissociation constant in water for a monorhamnolipid mixture extracted from Pseudomonas aeruginosa ATCC 9027 has been determined using potentiometry and two spectroscopic approaches at concentrations below and above the critical micelle concentration (cmc). Potentiometric titrations resulted in pKa values ranging from 4.28 +/- 0.16 to 5.50 +/- 0.06 depending on concentration. 1H NMR spectrochemical titrations at concentrations below the cmc revealed a pKa value of 4.39 +/- 0.06. ATR-FT-IR spectrochemical titrations on solutions well above the cmc gave a pKa value of 4.84 +/- 0.05. The value of 4.28 for the free rhamnolipid molecule for concentrations below the cmc differs markedly from that reported previously. However, the pKa of 5.50 for surface-adsorbed and solution aggregates correlates closely to that previously reported. Differences in these pKa values are rationalized in terms of the pH- and concentration-dependent aggregation behavior of rhamnolipids in aqueous solution.

Fatty Acid Cosubstrates Provide β-Oxidation Precursors for Rhamnolipid Biosynthesis in Pseudomonas aeruginosa, as Evidenced by Isotope Tracing and Gene Expression Assays

“…Pseudomonas aeruginosa strain ATCC 9027, which produces only monorhamnolipids (18,41), was purchased from the American Type Culture Collection (Rockville, MD) and stored as a glycerol freezer stock at Ϫ80°C. A peptone, yeast extract, glucose (PTYG) plate (Difco) was prepared from the freezer stock, and a single colony was inoculated into a preculture composed of NH 4 for 96 h with constant shaking at 200 rpm.…”

“…Rhamnolipids were harvested and purified after 96 h as described previously (18). Briefly, cells were separated from the supernatant by centrifugation at 10,000 rpm for 10 min, subjected to acid precipitation, extracted with a 9:1 ratio of chloroform to methanol, and concentrated by rotoevaporation.…”

“…Treatment effects of the carbon sources on the rhamnolipid yields were evaluated by one-way analysis of variance (ANOVA) using the Tukey-Kramer honestly significant difference (HSD) test (␣ ϭ 0.05). Significant differences between the means for the RER of each target gene of C 18 and C 18 -d 35 treatments were evaluated separately at each time point using a two-tailed t test (P Յ 0.05).…”

Formation and stabilization of nanoemulsions using biosurfactants: Rhamnolipids

“…1). Typically, the carboxylic acid groups of rhamnolipd have pK a values around pH 4.28-5.50 [50], which would account for the steep decrease in ζ-potential observed around these values (Fig. 8b).…”

Improved detection of rhamnolipid production using agar plates containing methylene blue and cetyl trimethylammonium bromide

“…The circles were not apparent at pH 3 and 5, because the ion-pairing requires rhamnolipids to be present as anionic ions while their pKa were in the range of 4.3-5.5 (depending on the rhamnolipid structure and concentration) (Lebron-Paler et al 2006). At pH 11, the circles were formed with comparable sizes to those at pH 7 and 9 but the agar plates at this pH were a much paler blue making the circle identification more difficult.…”