# Precipitation of Hemicelluloses from DMSO/Water Mixtures Using Carbon Dioxide as an Antisolvent

Authors: Emmerich Haimer; Martin Wendland; Antje Potthast; Thomas Rosenau; Falk Liebner
Journal: Journal of Nanomaterials
 (2008)

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## Abstract

Supercritical antisolvent precipitation is a relatively recent technology which can be used for controlled preparation of polymer particles from solutions. This is done by the addition of an antisolvent to a polymer solution causing supersaturation of the polymer, especially under supercritical conditions. The particle size of the precipitates can be adjusted mainly by the rate of supersaturation. Spherical xylan or mannan particles having a narrow particle size distribution were precipitated from hemicellulose solutions in dimethyl-sulfoxide (DMSO) or DMSO/water mixtures by carbon dioxide as an antisolvent. By depending on the type of hemicellulose, the DMSO/H2O ratio, and the precipitation conditions such as pressure and temperature, the resulting particle size can be adjusted within a wide range from less than 0.1 to more than 5 μm. Nano- and microstructured native xylans and mannans as obtained can be used in many applications such as encapsulation of active compounds, slow release agents, or chromatographic separation materials.


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## Body

## 1. Introduction

Biopolymers  as renewable raw materials are currently attracting enormous interest. Hemicelluloses  represent
an important percentage amongst these classes of  natural compounds. About 15–35% of the dry
biomass of woody tissues and up to 50% of cereal grains consist of such polysaccharides.
The most abundant hemicelluloses are xylans and mannans. Whereas hemicellulose
in hardwood mainly consists of pentosans (mostly xylans), the majority of
softwood hemicelluloses are hexosans with mannan as the dominating polysaccharide
[1].Apart from using hemicelluloses as feedstock for the production of chemicals,
their utilization for designing novel materials increasingly attracts interest
and is realizable but challenging as well. Applications which take advantage of
the unique properties of hemicelluloses are still more or less restricted to utilization
as natural adhesives in board and paper manufacturing. However, increasing
attention is paid to the bioside of these polymers. Xylans, for instance, are
able to regulate blood pressure, bind hydrophobic mutagens, inhibit
mutagenicity, and contribute to the effects of dietary fibers [2]. However,
purification of hemicelluloses is a crucial prerequisite to applications, in
particular on larger industrial scale.Several concepts of xylan purification have been proposed [3, 4]. Ebringerova
et al. [5] found that DMSO and DMSO/water mixtures are efficient solvents for
low-branched heteroxylans. An interesting feature of DMSO is its good
miscibility with CO2 at high pressure [6, 7]. Water can additionally
be used to modify the CO2/DMSO system for supercritical antisolvent
precipitation [8–10].For antisolvent precipitation, CO2 is mixed with a solution
of the respective solid in an organic solvent. The mixing process is indicated
as a straight line in the diagram of the ternary system DMSO/CO2/hemicellulose
(cf. Figure 1(b)). With
increasing CO2 content and after crossing the binodial, the
homogeneous solution splits into a solvent/CO2 phase (point A) and a
liquid phase which is rich in DMSO and hemicellulose (point A'). Continued addition
of CO2 to point D leads to supersaturation in the polymer-rich
liquid phase (point B') and finally precipitation of hemicellulose particles. A
scheme of the antisolvent precipitation process is given in Figure 1(a). After
precipitation, the solvent/CO2 phase can be flushed out, and remaining
traces of the solvent are finally removed from the precipitate by a continuous flow
of supercritical carbon dioxide (scCO2; supercritical drying).(a) Scheme of antisolvent
precipitation and (b) diagram of the ternary system DMSO/CO2/hemicellulose.
(a)(b)Temperature and pressure can be varied to influence the precipitation rate and, thus, particle
size and shape. Particle formation below the critical point of the mixture is
strongly influenced by the solvent-transfer resistance between the liquid and
gas phases.
Increasing pressure and temperature beyond the critical values of the solvent/antisolvent
mixture causes complete miscibility of the solvent with CO2. Precipitation
under these conditions is no longer influenced by any mass transfer resistance.In the present paper, we would like
to communicate the first results of antisolvent precipitation of hemicelluloses
from solutions containing DMSO as the principal solvent. The controlled
preparation of xylan and mannan particles was successful for both batch mode
and semicontinuous mode.


## 2. Experimental

All precipitation experiments were performed at 40°C. DMSO (>99.5%) was purchased
from Carl Roth GmbH & Co. KG, Karlsruhe, Germany and carbon dioxide (>99.8%) was
purchased from Linde (Austria).The studied xylan was extracted from beech
sulfite pulp (Lenzing AG, Austria),
and spruce mannan was kindly provided by S. Willför (Abo Academy, Turku, Finland).Pressure-dependent solubility of the hemicellulose
samples was determined by measuring the optical density of the solutions (OD 250 nm) at 10 different pressure levels covering the range from 1 to 150 bar. A
logistic dose response model was fitted to the data in order to relate the
reduced concentrationc/c0 to the applied pressure (Table Curve 2D
5.0, Cranes Software Inc, Troy, MI, USA).Batch experiments were performed using high-pressure equipment
consisting of an HPLC pump (TSP minipump, Thermal Separation Products, Toms River, NJ, USA) with cooled dual
pistons and an empty 40 mL semipreparative HPLC column (Alltech Grom GmbH, Rottenburg-Hailfingen,
Germany) with a stainless steel frit at the bottom as batch cell. 20 mL of the saturated
hemicellulose solutions (pure DMSO or DMSO/water mixtures) were placed in the batch
cell and heated up to the respective temperature. After temperature equilibration,
the column was pressurized with CO2 at a flow rate of 5 g min−1.
After 15 minutes of equilibration time, the supernatant liquor was flushed out with
scCO2. The precipitate was dried with scCO2 for 60 minutes.Semicontinuous experiments were performed in a 0.2 l
atomisation vessel (Separex, Champigneulles, France).
After pressurizing with CO2 and equilibrating  (5 minutes) the atomization
vessel at 150 bar and 40°C, the saturated polymer solution was sprayed
into the vessel at a flow rate of 1 mL min−1. Hemicellulose particles
were collected on a stainless steel frit at the bottom of the vessel. Removal
of the supernatant liquor and drying of the precipitate were carried out as described above. DMSO
can be recovered by depressurizing the supernatant.Scanning electron microscopy was performed on a Phillips XL 30 ESEM instrument.
All samples were sputtered with gold and mapped at an acceleration voltage of
10 kV.


## 3. Results and Discussion

The principle of antisolvent mediated precipitation of xylans and mannans from DMSO
solutions is based on a CO2-induced phase separation into a liquid phase
rich in DMSO and hemicelluloses and a solvent/antisolvent phase. With increasing
CO2 content, hemicellulose becomes augmented in the liquid phase (solvent
is transferred to the solvent/antisolvent phase) which eventually causes supersaturation
and precipitation of the hemicellulose. As solubility of the solute plays a
decisive role for precipitation, the pressure-dependent solubility of xylan and
mannan samples in the binary mixture DMSO/CO2 was studied starting
from saturated (1010 mbar, 25°C) solutions of the hemicelluloses in pure DMSO.The solubility of
xylan and mannan in DMSO becomes quite different when approaching the critical
pressure of the binary mixture DMSO/CO2 at 40°C (see Figure 2).Figure 2
Pressure dependence of the solubility of xylan and mannan in
DMSO/CO2 at 40°C.Beyond the critical pressure of 87 bar, DMSO is fully miscible with scCO2 and supersaturation occurs very fast. However, even under subcritical conditions a
certain amount of the solute can be precipitated. Thus, two different pathways exist
to obtain hemicellulose particles from solutions: supersaturation (a) below or (b) beyond
full miscibility of the antisolvent with the solvent. Below full miscibility,
the precipitation process is limited by the mass transfer of CO2 into DMSO, and supersaturation of the solute is reached only slowly, which
results in the formation of comparatively large hemicellulose particles. In
case of supercritical conditions, precipitation occurs very fast, and much
smaller particles can be obtained.In batch mode, where a solution of the particular hemicellulose in DMSO
is slowly pressurized with CO2, supersaturation at subcritical
conditions and hence size of the precipitating particles can be easily
controlled by CO2 flow rate and final pressure. At a low flow rate
of 5 g min−1 and at a pressure of 90 bar, particles with an average
diameter of 2–5 μm were obtained.
An important factor affecting the particle size is the type of hemicellulose. Whereas
spherical xylan particles having a diameter of 2-3 μm were obtained
from DMSO in batch mode, distinctly bigger particles (5-6 μm) were
obtained for mannan under the same conditions  (cf. Figure 3).(a) SEM
pictures of xylan and (b) mannan particles obtained in batch mode by precipitation from DMSO
solutions using carbon dioxide as antisolvent.
(a)(b)The SEM pictures also reveal the presence of very small particles which
might be formed due to impurities,
fragmentation of initially formed bigger particles, or partially fast precipitation
of a smaller hemicellulose fraction. The latter is most likely as supercritical
conditions were achieved at the end of the precipitation process in order to
initiate scCO2 drying.It is evident from the SEM pictures that all hemicellulose precipitates are low porous.
This can be explained by a liquid-liquid phase separation occurring during subcritical
precipitation after having overcome the mass transfer resistance of the binary solvent/antisolvent
system [11]. During this process, agglomeration of solute molecules and eventually
precipitation of polymer particles occur. Under subcritical conditions, the agglomerates usually form
droplets which can be stable or unstable upon drying with scCO2. Precipitation
of xylan under subcritical conditions, for example, yields stable droplets. In
contrast, mannan forms droplets that are only partly stable. As the particle
size of the mannan precipitate is distinctly larger, it might be better suited for applications like coating.The addition of small amounts of water shifts the critical point of the binary
system DMSO/CO2, and hence the coexistence of the two phases, to
higher pressure values [7, 8]. Upon addition of a sufficient amount of water, a
liquid-liquid phase split occurs in the ternary system as it is well known from
other ternary systems consisting of H2O, CO2, and any
polar organic solvent [12]. The formation of this additional phase increases the
above-mentioned mass transfer resistance, causes slower precipitation, larger
particle sizes, and enhanced agglomeration compared to water-free DMSO/CO2 mixtures.Increasing water contents also extend the subcritical range of the ternary
system  DMSO/H2O/CO2. This effect is quite advantageous
since it facilitates tuning of the particle size and morphology as the amount
of added water can be freely chosen
due to the insolubility of hemicelluloses in water. SEM pictures of
hemicellulose particles precipitated from aqueous DMSO (10%  H2O,
v/v) are shown in Figures 4(a) and 4(b).(a) SEM pictures of subcritically
precipitated xylan and (b) mannan obtained in batch mode from aqueous DMSO (10%H2O, v/v). (c) SEM picture of xylan precipitates obtained in
semicontinuous mode by spraying a solution of xylan in DMSO into scCO2 at 40°C and 150 bar.
(a)(b)(c)The size of the individual xylan particles—no matter whether
obtained by precipitation from pure or aqueous DMSO—was very similar.
Spheres of about 1μm were initially formed which agglomerated to stable, grape-like
clusters with an average diameter of about 10 μm. Mannan tends to form even
bigger clusters under the same precipitation conditions, and also sponge-like
structures can be obtained (cf.
Figure 4(b)).In contrast to the two batch methods where the principle of subcritical
precipitation is applied to obtain large hemicellulose particles from DMSO or
DMSO/H2O mixtures, very small particles are formed when xylan or
mannan is precipitated beyond the critical pressure of the binary system
DMSO/CO2 at 40°C (cf.
Figure 4(c)). This can be achieved by semicontinuously spraying a nearly
saturated hemicellulose solution in DMSO into scCO2 (e.g., 150 bar, 40°C).
As DMSO and CO2 are fully miscible under supercritical conditions,
mass transfer resistance becomes very low; supersaturation develops fast and
hence very small particles precipitate. For the studied xylans, particle diameters
in the 10 nm range were obtained.The results  obtained so far confirm the proposed mechanism of antisolvent
precipitation. As solvent and antisolvent are fully miscible under
supercritical conditions, supersaturation, solidification, and liquid-liquid
phase separation occur at the same time. Thus, no droplets are formed and the
resulting particles do not agglomerate.


## 4. Conclusion

Common chemical precipitation methods and thermal drying methods are often harmful to
thermolabile substances, as many biopolymers are. Antisolvent precipitation in
combination with supercritical drying is a valuable alternative for controlled
precipitation, adjusting of largely uniform particle sizes and retention of the
fragile particle structures.Antisolvent precipitation can be
performed in both batch and semicontinuous modes. In batch mode, low isothermal pressurization
rates afford precipitation under subcritical conditions. Semicontinuous
precipitation can be performed subcritically as well as supercritically.In batch mode, pressurizing rate and
type of hemicellulose largely affect the size and agglomeration tendency of the
precipitates. Low CO2 pressurizing rates were found to yield large
particles. The studied mannan had a higher agglomeration tendency than the
xylan.The addition of small quantities of
water extends the subcritical range of the ternary system DMSO/H2O/CO2.
This effect can be used to tune the particle size and morphology as the amount
of added water can be freely chosen
due to the insolubility of hemicelluloses in water. Batch precipitation in
presence of water can be used to prepare large xylan aggregates or sponge-like mannan
structures. Small particles with a very narrow particle size distribution can
be obtained in semicontinuous mode.Antisolvent precipitation facilitates
controlled preparation of hemicellulose particles at comparatively moderate
conditions (40°C, <100 bar) and enables a recovery of DMSO and CO2.
The described technology has a great potential for increasing the number of hemicellulose
applications.



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*Source: 826974-2009-01-05.xml*