Enhancing the Membranolytic Activity of Chenopodium quinoa Saponins by Fast Microwave Hydrolysis
"> Figure 1
<p>Alkaline hydrolysis of Saponin O to Saponin O<sup>h</sup>.</p> "> Figure 2
<p>Schematic structure of the saponin molecules extracted from the quinoa husk. All the functions in R<sub>1</sub>, R<sub>2</sub> and R<sub>3</sub> are described in <a href="#molecules-25-01731-t001" class="html-table">Table 1</a>.</p> "> Figure 3
<p>General structures of the sapogenins detected in <span class="html-italic">Chenopodium quinoa</span> extract in previous reports [<a href="#B6-molecules-25-01731" class="html-bibr">6</a>,<a href="#B28-molecules-25-01731" class="html-bibr">28</a>].</p> "> Figure 4
<p>Mass spectrometry analysis of the quinoa saponin extract: MALDI-MS (+) spectrum (DHB/DMA as the matrix) and schematic representation of the [2 + 1] and [3 + 1] saponin structures. Ions detected at <span class="html-italic">m</span>/<span class="html-italic">z</span> 791 and 833 are fragment ions from the intact saponin ions. This is demonstrated using LC-MS analysis as shown in the SI.</p> "> Figure 5
<p>Ion mobility experiments on quinoa saponin ions: Arrival Time Distributions (ATD) of protonated (<span class="html-italic">m</span>/<span class="html-italic">z</span> 973) and sodiated (<span class="html-italic">m</span>/<span class="html-italic">z</span> 995) Saponin B. The <sup>TW</sup>CCS<sub>N2⭢He</sub> presented at the top of the signals are calculated using the calibration procedure presented in the experimental section. For the instrumental conditions used in the ion mobility experiments, see the Experimental Section. The signals marked with (*,<sup>#</sup>) correspond to the doubly-charged [M + 2Na]<sup>2+</sup> ions and to a fragment of ionized saponin O–The loss of glucose at C28 upon ion activation from [Saponin O + Na]<sup>+</sup> generates [Saponin O<sup>h</sup> + Na]<sup>+</sup> (see <a href="#molecules-25-01731-f001" class="html-fig">Figure 1</a>) that is the monodesmosidic isomer of [Saponin B + Na]<sup>+</sup>.</p> "> Figure 6
<p>Molecular diversity of the three and four-sugar saponins from the quinoa extract. Integration of the MALDI-ToF, LC-MS(MS), and LC-IMS-MS data with (<b>a</b>) the retention times in LC (min) and (<b>b</b>) the collision cross sections in IMS (Å<sup>2</sup>). Section areas are associated with LC-MS semi-quantitative analysis using Hederacoside C as the internal standard and correspond to molar proportions (% in the quinoa husk extract). The relative proportions are also given in <a href="#molecules-25-01731-t001" class="html-table">Table 1</a>.</p> "> Figure 7
<p>Microwave-assisted hydrolysis of the quinoa bidesmosidic saponins (5 min at 150 °C): influence of the pH on the extent of the consecutive hydrolysis reactions from Saponin B as determined using LC-MS. The given <span class="html-italic">m</span>/<span class="html-italic">z</span> ratios correspond to the [M + Na]<sup>+</sup> ions, but for the estimation of the relative proportions, both the [M + H]<sup>+</sup> and the [M + Na]<sup>+</sup> ions have been considered.</p> "> Figure 8
<p>Microwave-assisted alkaline hydrolysis (5min, pH 10, 150 °C) of Chenopodium quinoa saponin extract: MALDI (+) mass spectra of (<b>a</b>) the saponin extract and (<b>b</b>) the hydrolyzed saponins.</p> "> Figure 9
<p>Microwave-assisted hydrolysis of the quinoa bidesmosidic saponins (5 min at pH 10): influence of the temperature (60,90,120,150,180 °C) on hydrolysis reactions from quinoa husk saponin determined by MALDI-MS.</p> "> Figure 10
<p>Microwave-assisted alkaline hydrolysis (5min, pH 10, 150 °C) of Chenopodium quinoa saponin extract: composition of the hydrolysate. Integration of the MALDI-ToF, LC-MS(MS), and LC-IMS-MS data with (<b>a</b>) the retention times in LC (min) and (<b>b</b>) the collision cross sections in IMS (Å<sup>2</sup>). Section areas are associated with LC-MS semi-quantitative analysis using Hederacoside C as the internal standard and correspond to molar proportions (% in the hydrolyzed quinoa husk extract). The relative proportions are also given in <a href="#molecules-25-01731-t002" class="html-table">Table 2</a>.</p> "> Figure 11
<p>Schematic structure of the hydrolyzed saponins extracted from the quinoa husk. All the functions in R<sub>1</sub>, R<sub>2</sub> and R<sub>3</sub> are described in <a href="#molecules-25-01731-t002" class="html-table">Table 2</a>.</p> "> Figure 12
<p>Estimation of the membranolytic property of the natural and hydrolyzed saponins via the hemolytic activity assay. Monitoring the free heme absorbance (540 nm) with regards to the increasing saponin concentration. Hydrolyzed saponins appear to be more cytotoxic than the non-hydrolyzed ones. Hemolytic activity experiments were performed in triplicates and the average data as well as the standard deviations are gathered in <a href="#app1-molecules-25-01731" class="html-app">Table S1</a>.</p> "> Figure 13
<p>IMPALA simulation of Saponin O (dark line) and Saponin O<sup>h</sup> (grey line) traversing a 36 Å thick implicit membrane. The “energy-like” profile of the saponin traversing the implicit bilayer. The Z-axis corresponds to the position of the center of mass of the saponin along an axis orthogonal to the membrane, the center of the bilayer corresponding to the intersection with the Y-axis. The different circles represent the two most stable positions of both molecules the center of the bilayer corresponds to the Y-axis with the red circles corresponding to the most stable positions. The vertical lines represent the water/membrane interface (pink), the hydrophilic head/hydrophobic tail interface of the phospholipid bilayer (purple), and the center of the bilayer (yellow).</p> ">
Abstract
:1. Introduction
2. Results and Discussion
2.1. Saponin Identification and Quantification in the Quinoa Extract (QE)
2.2. Selectivity of the Microwave-Assisted Hydrolysis of Saponins
2.3. Activity Modulation–Hemolytic Activity Assay
2.4. Activity Modulation–In Silico Evaluation of the Mono- vs. Bidesmosidic Saponin Activities
3. Conclusions
4. Materials and Methods
Chemicals, Plant Sampling and Saponin Extractions
Chemicals
Saponin Extraction from Quinoa Husks
Microwave Hydrolysis
Mass Spectrometry Analysis
LC-MS Analyses
Semi-Quantitative Analysis
Hemolytic Activity
In silico Study
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Sample Availability: Samples of the compounds are not available from the authors. |
Composition | m/z [M + Na]+ | Δ(m/z) (ppm) | R1 | R2 | Aglycone | R3 | RT (min) | CCS (Ų) [M + Na]+ | CCS (Ų) [M + H]+ | Molar Proportion(%) | Mass Fraction (mg·g−1) | |
---|---|---|---|---|---|---|---|---|---|---|---|---|
I | C47H76O18 | 951.4929 | 2.1 | - CH3 | - CH2OH | Hed | Glc – Ara - | 7.3 | 234 | 254 | 14.9 | 6.1 |
Unknown-1 | C47H74O19 | 965.4722 | 0.2 | ? | ? | ? | Glc – Ara - | 5.3 | 235 | 254 | 1.9 | 0.8 |
19 | C47H76O19 | 967.4878 | 4.9 | - CH2OH | - CH2OH | AG489 | Xyl - Glc - | 4.0 | 238 | 254 | 1.6 | 0.7 |
19a | - CH2OH | - CH2OH | AG489 | Xyl - Glc - | 4.8 | 238 | 256 | 0.3 | 0.1 | |||
H | C48H76O19 | 979.4878 | 1.5 | - COOCH3 | - CH3 | SA | Glc – Ara - | 7.1 | 235 | NA | 1.6 | 0.7 |
70 | - CH3 | - CH3 | OA | Glc – GlcA - | 7.5 | 235 | NA | 0.9 | 0.4 | |||
Q | C48H78O19 | 981,5035 | 0.8 | - CH3 | - CH2OH | Hed | Glc – Gal - | 6.5 | 235 | 260 | 0.2 | 0.1 |
B | C48H76O20 | 995.4828 | 3.2 | - COOCH3 | - CH2OH | PA | Glc – Ara - | 4.9 | 243 | 260 | 61.7 | 25.3 |
61 | C53H86O23 | 1113.5458 | 1.6 | - CH3 | - CH2OH | Hed | Glc – Glc – Ara - | 5.9 | 275 | 290 | 0.7 | 0.3 |
Unknown-2 | C53H86O24 | 1127.5251 | 2.1 | ? | ? | ? | Glc – Glc – Ara - | 4.5 | 270 | 290 | 1.7 | 0.7 |
G | C54H86O24 | 1141.5407 | 3.6 | - COOCH3 | - CH3 | SA | Glc – Glc – Ara - | 5.7 | 266 | 291 | 3.0 | 4.6 |
Composition | m/z [M + Na]+ | Δ(m/z) (ppm) | R1 | R2 | Aglycone | R3 | RT (min) | CCS (Ų) [M + Na]+ | CCS (Ų) [M + H]+ | Molar Proportion (%) | |
---|---|---|---|---|---|---|---|---|---|---|---|
Ih | C41H66O13 | 789.4401 | 0.8 | - CH3 | - CH2OH | Hed | Glc – Ara - | 10.7 | 224 | 219 | 16.5 |
Unknown-1h | C41H64O14 | 803.4194 | 0.4 | ? | ? | ? | Glc – Ara - | 7.2 | 225 | 221 | 2.4 |
19h | C41H66O14 | 805.4350 | 0.7 | - CH2OH | - CH2OH | AG489 | Xyl - Glc - | 5.6 | 227 | 221 | 2.2 |
19ah | - CH2OH | - CH2OH | AG489 | Xyl - Glc - | 5.8 | NA | NA | 0.1 | |||
Hh | C42H66O14 | 817.4350 | 2.1 | - COOCH3 | - CH3 | SA | Glc – Ara - | 9.4 | 240 | NA | 2.2 |
70h | - CH3 | - CH3 | OA | Glc – GlcA - | 13.1 | 235 | NA | 1.0 | |||
Qh | C42H68O14 | 819.4501 | 0.6 | - CH3 | - CH2OH | Hed | Glc – Gal - | 9.4 | 228 | 222 | 0.1 |
Bh | C42H66O15 | 833.4299 | 1.1 | - COOCH3 | - CH2OH | PA | Glc – Ara - | 9.9 | 234 | 227 | 61.4 |
61h | C47H76O18 | 951.4929 | 1.3 | - CH3 | - CH2OH | Hed | Glc – Glc – Ara - | 10.2 | 250 | 253 | 0.4 |
Unknown-2h | C47H74O19 | 965.4722 | 2.5 | ? | ? | ? | Glc – Glc – Ara - | 7.1 | 252 | 254 | 0.9 |
Gh | C48H76O19 | 979.4879 | 0.5 | - COOCH3 | - CH3 | SA | Glc – Glc – Ara - | 9.4 | 265 | NA | 2.7 |
Oh | C48H76O20 | 995.4828 | 0.4 | - COOCH3 | - CH2OH | PA | Glc – Glc – Ara - | 7.6 | 258 | 263 | 10.1 |
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Colson, E.; Savarino, P.; J.S. Claereboudt, E.; Cabrera-Barjas, G.; Deleu, M.; Lins, L.; Eeckhaut, I.; Flammang, P.; Gerbaux, P. Enhancing the Membranolytic Activity of Chenopodium quinoa Saponins by Fast Microwave Hydrolysis. Molecules 2020, 25, 1731. https://doi.org/10.3390/molecules25071731
Colson E, Savarino P, J.S. Claereboudt E, Cabrera-Barjas G, Deleu M, Lins L, Eeckhaut I, Flammang P, Gerbaux P. Enhancing the Membranolytic Activity of Chenopodium quinoa Saponins by Fast Microwave Hydrolysis. Molecules. 2020; 25(7):1731. https://doi.org/10.3390/molecules25071731
Chicago/Turabian StyleColson, Emmanuel, Philippe Savarino, Emily J.S. Claereboudt, Gustavo Cabrera-Barjas, Magali Deleu, Laurence Lins, Igor Eeckhaut, Patrick Flammang, and Pascal Gerbaux. 2020. "Enhancing the Membranolytic Activity of Chenopodium quinoa Saponins by Fast Microwave Hydrolysis" Molecules 25, no. 7: 1731. https://doi.org/10.3390/molecules25071731