Production, characterization and biomedical potential of biosurfactants produced by haloalkaliphilic archaea from Wadi El-Natrun, Egypt

Abstract

Extreme halophilic archaea that can live in high saline environments can offer potential applications in different biotechnological fields. This study delves into the fascinating field of halophilic archaea and their ability to produce biosurfactants. Some strains of haloarchaea were isolated from Wadi El-Natrun and were screened for biosurfactants production in a standard basal medium using emulsification index assay. Two strains were chosen as the potential strains for surface tension reduction. They were identified as Natrialba sp. BG1 and N3. The biosurfactants production was optimized and the produced emulsifiers were partially purified and identified using FTIR and NMR. Sequential statistical optimization, Plackett-Burman (PB) and Box-Behnken Designs (BBD) were carried out using 5 factors: oil, NaCl, casamino acids, pH, and inoculum size. The most significant factors were used for the next Response Surface Methodology experiment. The final optimal conditions for biosurfactants production were the inoculum size 2% pH 11 and NaCl 250 g/L, for Natrialba sp. BG1 and inoculum size 2.2%, pH 10 and NaCl 100 g/L for Natrialba sp. N3. The produced biosurfactants were tested for wound healing and the results indicated that Natrialba sp. BG1 biosurfactants is more efficient than Natrialba sp. N3 biosurfactants. Biosurfactants extracts were tested for their cytotoxic effects on normal cell line as well as on different cancer cells using MTT assay. The findings demonstrated that varying concentrations of the biosurfactants (31.25, 62.5, 125, 250, 500 and 1000 µg/mL) exhibited cytotoxic effects on the cell lines being tested. Additionally, the outcomes unveiled the presence of anti-inflammatory and antioxidant properties for both biosurfactants. Consequently, they could potentially serve as natural, safe, and efficient novel agents for combating cancer, promoting wound healing, and providing anti-inflammatory and antioxidant benefits.

Keywords: Anti-inflammatory; Anticancer; Biosurfactants; Halophilic archaea; Wound healing.

Fig. 1

Microscopic examination of the two potential isolates. Above: Natrialba sp. BG1 and down: Natrialba sp. N3 where A and E are Gram stain reaction. SEM micrographs of BG1: B and C with magnification X10,000, and X15,000, respectively. SEM micrographs of N3: E and F with magnification X10,000 and X15,000

Fig. 2

Main effect of the factors influencing the biosurfactants production by Natrialba sp. BG1 and N3 based on PBD

Fig. 3

Three-dimensional surface showing the relationships between the tested factors and the biosurfactants as response in the form of Emulsification index E24% produced by Natrialba sp. BG1 and N3 A showed that at high inoculum size and middle pH gave the highest biosurfactants production level. B showed that at high inoculum size value and middle value of NaCl gave the highest biosurfactants production level. C showed foci for maximum level of biosurfactants production at middle both NaCl and pH for Natrialba sp. BG1 also D indicated that at high inoculum size value and middle pH values the maximum production was achieved. E showed the maximum response at high inoculum size and middle value of NaCl (F) showed foci for maximum level of biosurfactants production at middle values of both NaCl and pH

Fig. 4

FT-IR Spectra for Natrialba sp. BG1 biosurfactants (A) and Natrialba sp. N3 biosurfactants (B)

Fig. 5

A C13 NMR for Natrialba sp. BG1 biosurfactants B 1H NMR for Natrialba sp. BG1 biosurfactants, C C13 NMR for Natrialba sp. N3 biosurfactants and D 1H NMR for Natrialba sp. N3 biosurfactants

Fig. 6

Wound closure % of Natrialba sp. N3 and BG1 biosurfactants

Fig. 7

Effect of biosurfactants produced by Natrialba sp. BG1 (A, B, C & D) on MG63, MCF7, PC3 and MG63 cells at different concentration respectively, and (E, F, G & H) showed effect of Natrialba sp. BG1 biosurfactants on MG63, MCF7, PC3 and MG63 cells at different concentration respectively

Fig. 8

Toxicity of the biosurfactants produced by Natrialba sp. BG1 and N3 on different cell lines (A) PC3, (B) MFC3, (C) MG63 and (D) Caco2

Fig. 9

Anti-inflammatory effect of Natrialba sp. N3 and BG1 biosurfactants using LPS

Fig. 10

Second order equation to calculate the equivalent antioxidant amount percentage in both Natrialba sp. BG1 and N3 biosurfactants