Name: pretreatment of lignocellulosic biomass with low-cost ionic liquids
Uploaded: 2016-08-10T13:00:00
Description: Watch this Scientific Journal Video about Pretreatment of Lignocellulosic Biomass with Low-cost Ionic Liquids at JoVE.com

The authors acknowledge the Grantham Institute for Climate Change and the Environment, Climate-KIC and EPSRC (EP/K038648/1 and EP/K014676/1) for funding and Pierre Bouvier for providing experimental data for pine pretreatments.

Note: The protic ionic liquids used in the process are synthesized in our laboratory, although some might be or become commercially available. The resulting ionic liquids are acidic and corrosive and probably skin/eye irritants (depending on the amine used), and must therefore be handled with care wearing appropriate PPE (lab coat, safety specs, resistant gloves).
1. Preparation
<ol>
	<li>Preparing and storing the lignocellulosic biomass
	<ol>
		<li>Obtain the lignocellulosi...

<ol>
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Microbiol. Biotechnol. 82 (5), 815-827 (2009).</li><li>Brandt, A., Gr&#228;svik, J., Hallett, J. P., Welton, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deconstruction+of+lignocellulosic+biomass+with+ionic+liquids.">Deconstruction of lignocellulosic biomass with ionic liquids.</a> Green Chem. 15, 550 (2012).</li><li>Chen, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inexpensive+ionic+liquids:[HSO+4]&#8722;-based+solvent+production+at+bulk+scale).">Inexpensive ionic liquids:[HSO 4]&#8722;-based solvent production at bulk scale).</a> Green Chem. 16 (6), 3098-3106 (2014).</li><li>Brandt, A., Chen, L., van Dongen, B. E., Welton, T., Hallett, J. 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G., Baker, S. R., Decker, <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NREL/TP-5100-63351.">NREL/TP-5100-63351.</a> Low Solids Enzymatic Saccharificatin of Lignocellulosic Biomass. , (2015).</li><li>Brandt, A., Ray, M. J., To, T. Q., Leak, D. J., Murphy, R. 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The technique for the fractionation of lignocellulosic biomass presented here produces a cellulose-rich pulp and a lignin. Most of the hemicelluloses are dissolved into the ionic liquid and hydrolyzed, but not recovered. If hemicellulose sugars are desired, a hemicellulose pre-extraction step prior to the Ionosolv delignification may be necessary. It has so far been impossible to fully close the mass balance for the biomass, as it is not possible to identify and quantify all degradation products found in the ionic liquid...

Meeting humanity&#39;s energy demand sustainably is one of the greatest challenges that our civilization faces. Energy use is predicted to double in the next 50 years, putting greater strain on fossil fuel resources. 1 The buildup of greenhouse gases (GHG) in the atmosphere through wide-spread fossil fuel use is particularly problematic, as CO2 generated from combustion of fossil fuels is responsible for 50% of the anthropogenic greenhouse effect. 2 Therefore, large-scale application of renewable and carbon neutral technologies is essential for meeting the increased energy and material needs of future generations. 1, 3
Plant biomass is the most versatile renewable resource, as it can be used to produce heat, electricity as well as carbon-based chemicals, materials and fuels. Primary advantages of lignocellulosic biomass over other biomass types are its abundance, potential for high yields per area of land and often much higher CO2 emission savings, which includes high retention of carbon in the soil. 4, 5 Additional benefits of using biomass include local availability, low capital requirements to convert biomass to energy, and soil erosion prevention. 8
Major producers of lignocellulosic feedstocks are the forestry industry and the agricultural sector as well as municipal waste management. 6 Lignocellulose production has the potential to be expanded, with a mind to limiting deforestation and avoiding the replacement of food crops and release of potential pollutants. 7 For renewable biomass to become a viable widespread source of liquid transportation fuels and chemicals, its processing must become economically competitive with fossil fuel conversion technologies. 9, 10 A key to achieving this is to boost the yield and quality of biomass-derived intermediates while reducing cost.
Lignocellulose contains a high proportion of sugars which can be converted to fuels and chemicals via catalytic and microbial conversions. 11 These sugars are present in lignocellulose in polymeric form as cellulose and hemicellulose. They can be hydrolyzed into glucose and other sugar monomers and then used for producing bioethanol and other bio-derived chemicals and solvents. 12
In order to access the cellulosic sugars, pretreatment of the biomass is necessary through physical, chemical, or combined processes. 4 The pretreatment is arguably the most costly step in the valorization of lignocellulosic biomass. Hence research into improved pretreatment processes is imperative.
Various pretreatment technologies are available. Of particular interest are those that separate the lignin from cellulose (fractionative pretreatment). Lignin, the third major component in lignocellulose, limits access of hydrolyzing agents to cellulose and hemicellulose and reduces the sugar yield per ton of feedstock. 11 The separated lignin can be utilized as an additional biorefinery intermediate if it is isolated in suitable quality. 13 One fractionative process is the Kraft process which is the most common pretreatment for paper/cellulose production. In Kraft pulping, wood chips are placed in a mixture of sodium hydroxide and sodium sulfide and heated at elevated temperatures of around 170 &#176;C under high pressure. 14 The alkaline reactions remove hemicellulose and lignin by breaking the polymers down to short fragments via nucleophilic and base catalysis, and by dissolving the lignin fragments via de-protonation of phenolic hydroxyl/alcohol groups. Another common delignification process is the Organosolv process which also fragments and dissolves the lignin and hemicellulose. Rather than using an alkaline aqueous solution, organic solvents such as ethanol and acetic acid are used at high temperatures ranging between 160-200 &#176;C and pressures from 5-30 bar. Organosolv pretreatment has some advantages over Kraft pulping in that it produces less air and water pollution. 15 Both processes possess some economic challenges, if used for production of chemicals and fuels rather than cellulose. 16 The Ionosolv pretreatment uses ionic liquids, which are salts that have melting points below 100 &#176;C and, as a result of their powerful Coulombic interactions, very low vapor pressures. 17 This eliminates air pollution in the pretreatment process, and enables processing at or near atmospheric pressure.
While most ILs are created in laborious, multi-step syntheses, protic ILs can be synthesized in a one-step process from commodity chemicals, which makes them less expensive; it is estimated that some ILs could be produced at bulk scale for a price of $1.24 per kg which is comparable to common organic solvents such as acetone and toluene. 18 The ability to recycle and reuse these customizable ILs in a process that operates at comparatively lower temperatures and pressures makes this a more benign alternative and an economically attractive candidate for biorefining.
This detailed video protocol demonstrates a lab-scale version of the Ionosolv process for the delignification of lignocellulosic biomass and the eventual enzymatic saccharification of the cellulose-rich pulp as well as the recovery of a high-purity odor-free lignin.19

<table border="0" cellpadding="0" cellspacing="0" width="851">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">IL synthesis</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td style="width:267px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Round bottom flask, with standard ground joint 24/29 NS, 1000 ml</td>
			<td style="width:116px;">Lenz</td>
			<td style="width:131px;">3 0024 70</td>
			<td>VWR product code 271-1309&#160;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">250mL Addition Funnel, Graduated, 29/26 Joint Size, 0-4mm PTFE Valve</td>
			<td style="width:116px;">GPE</td>
			<td style="width:131px;">CG-1714-16</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Dish-shaped dewar flask, SCH 31 CAL&#160;</td>
			<td style="width:116px;">KGW-Isotherm</td>
			<td style="width:131px;">1197</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Volumetric flask, 200 ml</td>
			<td style="width:116px;">VWR</td>
			<td style="width:131px;">612-3745&#160;</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Cork rings, pasteur pipettes and teet, wash bottle with deionised water, large magentic stir bar</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Biomass size reduction</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Heavy Duty Cutting Mill SM2000&#160;</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td>Discontinued</td>
			<td style="width:267px;">Replaced with Cutting Mill SM 200 (20.728.0001)&#160;</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Bottom sieves (10 mesh square holes, for particle size &#60;2 mm)</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td style="width:131px;">03.647.0318</td>
			<td style="width:267px;">Part of cutting mill</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Analytical Sieve Shaker AS 200</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td>30.018.0001</td>
			<td>Part of sieving machine</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Test Sieve 200 mm &#216; x 50 mm height ISO 3310/1 (180 &#181;m)</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td>60.131.000180</td>
			<td>Part of sieving machine</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Test Sieve 200 mm &#216; x 50 mm height ISO 3310/1 (850 &#181;m)</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td style="width:131px;">60.131.000850</td>
			<td align="left">Part of sieving machine</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Collecting pan, stainless steel, 200 mm &#216;, height 50 mm&#160;</td>
			<td style="width:116px;">Retsch&#160;</td>
			<td style="width:131px;">69.720.0050</td>
			<td>Part of sieving machine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Rotary evaporator:</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Rotary evaporator (Rotavapor R-210)</td>
			<td style="width:116px;">Buchi&#160;</td>
			<td>Discontinued</td>
			<td>Replaced with Rotavapor R-300</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Water bath (Heating bath B-491)</td>
			<td style="width:116px;">Buchi&#160;</td>
			<td>48201</td>
			<td>Part of rotary evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Recirculator&#160;</td>
			<td style="width:116px;">Julabo</td>
			<td style="width:131px;">F25</td>
			<td>Part of rotary evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum pump (MPC 101 Z)</td>
			<td style="width:116px;">Ilmvac GmbH</td>
			<td style="width:131px;">412522</td>
			<td>Part of rotary evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum controller (Vacuum Control Box VCB 521)</td>
			<td style="width:116px;">Ilmvac GmbH</td>
			<td style="width:131px;">600053</td>
			<td>Part of rotary evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Parallel evaporator:</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">StarFish Base Plate 135mm (for Radleys &#38; IKA)&#160;</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95010</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Monoblock for 5 x 250ml Flasks</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95130&#160;</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Telescopic 5-way Clamp with Velcro</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95400</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Gas/Vacuum Manifold with connectors</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95510&#160;</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">650mm Rod</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95665&#160;</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Quick Release Male, R/A Barbed 6.4mm + Shut-off (3.2mm ID)&#160;</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95520</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Stirrer/hot plate</td>
			<td>Radleys</td>
			<td>RR98072</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Temperature controller</td>
			<td style="width:116px;">Radleys</td>
			<td>RR98073</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Elliptical Stirring Bar 15mm Rare Earth</td>
			<td style="width:116px;">Radleys</td>
			<td>RR98097&#160;</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum cold trap, plastic coated, PTFE stopcock</td>
			<td style="width:116px;">Chemglass</td>
			<td style="width:131px;">CG-4519-01</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum pump (MPC 101 Z)</td>
			<td style="width:116px;">Ilmvac GmbH</td>
			<td style="width:131px;">412522</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Tygon tubing E-3603, 6,40 mm (internal) 12,80 mm (external)&#160;&#160;</td>
			<td style="width:116px;">Saint-Gobain/VWR</td>
			<td style="width:131px;">228-1292&#160;</td>
			<td>Part of parallel evaporator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Parallel Soxhlet extractor:</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">StarFish Base Plate 135mm (for Radleys &#38; IKA)</td>
			<td>Radleys</td>
			<td>RR95010&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Monoblock for 5 x 250ml Flasks</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95130&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Telescopic 5-way Clamp with Velcro</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95400&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Telescopic 5-way Clamp with Silicone Strap and Long Handle</td>
			<td>Radleys</td>
			<td>RR95410&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Water Manifold with connectors</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95500&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">650mm Rod</td>
			<td style="width:116px;">Radleys</td>
			<td>RR95665&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Quick Release Male, R/A Barbed 6.4mm + Shut-off (3.2mm ID)</td>
			<td>Radleys</td>
			<td>RR95520&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Coil condensers with standard ground joints 29/32 NS</td>
			<td>Lenz</td>
			<td>5.2503.04&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Extractor Soxhlet 40mL borosilicate glass 29/32 socket 24/29 cone</td>
			<td>Quickfit</td>
			<td>EX5/43&#160;</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Stirrer/hot plate</td>
			<td>Radleys</td>
			<td>RR98072</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Temperature controller</td>
			<td style="width:116px;">Radleys</td>
			<td>RR98073</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Recirculator</td>
			<td>Grant</td>
			<td>LTC1</td>
			<td>Part of soxhlet extractor</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Cellulose extraction thimble</td>
			<td>Whatman</td>
			<td>2280-228</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Tweezers</td>
			<td>Excelta</td>
			<td>20A-S-SE</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum drying oven:</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum drying oven</td>
			<td>Binder</td>
			<td>VD 23</td>
			<td>Part of vacuum oven</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Dewar vessel 2L 100x290mm with handle</td>
			<td>KGW-Isotherm</td>
			<td>10613</td>
			<td>Part of vacuum oven</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Vacuum Trap</td>
			<td>GPE</td>
			<td>CG-4532-01&#160;</td>
			<td>Part of vacuum oven</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Other equipment:</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Analytical balance</td>
			<td style="width:116px;">A&#38;D</td>
			<td style="width:131px;">GH-252</td>
			<td>accuracy to &#177; 0.1 mg</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Volumetric Karl Fischer titrator</td>
			<td style="width:116px;">Mettler Toledo</td>
			<td style="width:131px;">V20</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">10 mL disposable pipette</td>
			<td style="width:116px;">Corning Inc</td>
			<td style="width:131px;">Costar 4101 10 mL Stripette</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Eppendorf&#160;Research&#160;plus pipette, variable volume, volume 100-1000&#160;&#956;L</td>
			<td style="width:116px;">Eppendorf</td>
			<td>3120000062</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Desiccator</td>
			<td style="width:116px;">Jencons</td>
			<td style="width:131px;">JENC250-028BOM</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Ace pressure tube bushing type, Front seal, volume 15&#160;mL&#160;</td>
			<td style="width:116px;">Ace Glass</td>
			<td>8648-04&#160;</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Ace O-rings, silicone, 2.6&#160;mm, I.D. 9.2&#160;mm&#160;</td>
			<td style="width:116px;">Ace Glass</td>
			<td>7855216</td>
			<td>O-ring for pressure tube</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Vortex shaker</td>
			<td style="width:116px;">VWR International</td>
			<td style="width:131px;">444-1378 (UK)</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Fan-assisted convection oven</td>
			<td style="width:116px;">ThermoScientific</td>
			<td style="width:131px;">HeraTherm OMH60</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Oven glove (Crusader Flex)</td>
			<td style="width:116px;">Ansel Edmont</td>
			<td>42-325</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">250 mL Round bottom flask single neck ground joint 24/29 (Pyrex)</td>
			<td style="width:116px;">Quickfit&#160;</td>
			<td style="width:131px;">FR250/3S</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Rotaflo stopcock adapter with cone 24/29</td>
			<td style="width:116px;">Rotaflo England</td>
			<td style="width:131px;">MF11/2/SC</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">50 mL Falcon&#160; tube</td>
			<td style="width:116px;">Heraeus/Kendro</td>
			<td style="width:131px;">HERA 76002844</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Centrifuge (Mega Star 3.0)</td>
			<td style="width:116px;">VWR&#160;</td>
			<td style="width:131px;">521-1751</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Reagents:</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Ethanol absolute</td>
			<td style="width:116px;">VWR</td>
			<td style="width:131px;">20820.464</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Triethylamine</td>
			<td style="width:116px;">Sigma-Aldrich</td>
			<td style="width:131px;">T0886</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:339px;">Sulfuric acid 5 mol/l (10N) AVS TITRINORM volumetric solution Safe-break bottle 2,5L</td>
			<td>VWR</td>
			<td>191665V</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Purified water (15 M&#937; ressitance)</td>
			<td style="width:116px;">Elga</td>
			<td style="width:131px;">CENTRA R200</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Lignocellulosic biomass:</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Miscanthus X gigantheus</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:339px;">Pinus sylvestris</td>
			<td style="width:116px;"></td>
			<td style="width:131px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

pretreatment of lignocellulosic biomass with low-cost ionic liquids

A number of ionic liquids (ILs) with economically attractive production costs have recently received growing interest as media for the delignification of a variety of lignocellulosic feedstocks. Here we demonstrate the use of these low-cost protic ILs in the deconstruction of lignocellulosic biomass (Ionosolv pretreatment), yielding cellulose and a purified lignin. In the most generic process, the protic ionic liquid is synthesized by accurate combination of aqueous acid and amine base. The water content is adjusted subsequently. For the delignification, the biomass is placed into a vessel with IL solution at elevated temperatures to dissolve the lignin and hemicellulose, leaving a cellulose-rich pulp ready for saccharification (hydrolysis to fermentable sugars). The lignin is later precipitated from the IL by the addition of water and recovered as a solid. The removal of the added water regenerates the ionic liquid, which can be reused multiple times. This protocol is useful to investigate the significant potential of protic ILs for use in commercial biomass pretreatment/lignin fractionation for producing biofuels or renewable chemicals and materials.

The exact amount of lignin removal and lignin precipitation, recovered pulp and glucose yield depend on the type of biomass used, the temperature at which the treatment is run and the duration of the treatment. Short pretreatment times and low temperatures lead to incomplete pretreatment while at higher temperatures the cellulose becomes unstable in the ionic liquid, leading to hydrolysis and degradation. The selected ionic liquid also plays an important role in the outcome of the fractio...

Watch this Scientific Journal Video about Pretreatment of Lignocellulosic Biomass with Low-cost Ionic Liquids at JoVE.com

Pretreatment of Lignocellulosic Biomass with Low-cost Ionic Liquids

The pretreatment of lignocellulosic biomass with protic low-cost ionic liquids is shown, resulting in a delignified cellulose-rich pulp and a purified lignin. The pulp gives rise to high glucose yields after enzymatic saccharification.

A number of ionic liquids (ILs) with economically attractive production costs have recently received growing interest as media for the delignification of ...

The overall goal of this experimental method is to use low-cost ionic liquids for isolating lignin and cellulose from lignocellulosic biomass and understand factors affecting the separation and isolation of these biopolymers. This method can help answer key questions in the biorefinery field, such as how to isolate cellulose and a novel lignin using low-cost ionic liquids. Our technique utilizes unique solvent properties of ionic liquids, but our advantage is that these ionic liquids can be easily synthesized using low-cost starting materials.Visual demonstration of this method is critical as there are numerous steps that are a lot easier to learn and carry out once you've seen it, rather than reading a page-long description. Demonstrating this procedure will be Florence, Clem, Wei-Chien, and Lisa. All of whom are graduate students in my laboratory.Determine the water content of the synthesized or purchased ionic liquid solution by volumetric Karl Fischer titration according to instructions issued by the titrator manufacturer. Add a few drops of the ionic liquid into the titrator using a pre-weighed syringe. Enter the weight of liquid added into the titrator and wait until the titrator displays a reading.Record the water content. Reduce the water content to five weight percent below the desired water content using a rotary evaporator, and confirm the new water content by Karl Fischer titration as before. Use 10 grams of ionic liquid solution containing 20 weight percent water and a biomass to solvent ratio of one to 10, and perform the necessary experimental calculations as described in the text protocol.Pre-weigh three 15 milliliter pressure tubes with Teflon caps and silicone O-rings. Visually inspect the pressure tubes to ensure they have no cracks or flaws. With a 10 milliliter pipette, add the required amount of ionic liquid solution into the pressure tube standing on the scale.Use cork rings to keep the tube standing up. Record the weight of the ionic liquid solution added. Then add the calculated amount of water to the solution using a pipette, assuming the density of water to be one gram per milliliter.Add the required amount of air-dried liquid lignocellulose by placing a piece of aluminum foil on a balance, tearing the balance, and weighing out the biomass. Tear the balance again and add the biomass to the tube. Place the empty foil back on the balance and record difference.Close the lid using a Teflon cap with the silicone O-ring. Check for a good seal without over-tightening. Record the weight of the pressure tubes containing the biomass and ionic liquid.Then mix the contents of the tube by using a vortex shaker until all of the biomass is in contact with the ionic liquid. Place the pressure tubes in a fan-assisted oven that has been preheated to the desired temperature. For example, leave the tubes for eight hours at 120 degrees Celsius or for one hour at 150 degrees Celsius.Remove the vials from the oven using an oven glove, and place them in a fume hood to let them cool to room temperature. Check the weight of the vials after cooling to assure no water escaped during cooking. Transfer the contents of each tube into a 50 milliliter centrifuge tube using 40 milliliters of absolute ethanol.Shake the tube using a vortex shaker for one minute to mix well, and leave the tube at room temperature for at least one hour. After vortexing for another 30 seconds, centrifuge the tube for 50 minutes at 2, 000 times G.Separate the liquid and solid by careful decanting. Collect the liquid in a clean 250 milliliter round bottom flask with a stir bar.Add 40 milliliters of fresh ethanol into the 15 milliliter falcon tube and repeat the mixing and centrifugation step three times. Remove the ethanol from the ionic liquid by placing the round bottom flasks on a heating block. Connect each flask to a vacuum pump with a cold trap.Fill the trap with dry ice and set the heating to 40 degrees Celsius. Then switch on the stirring and the pump. Transfer the wet-washed pulp into cellulose thimbles and label each thimble using a pencil.Fill 150 milliliters of absolute ethanol into a clean 250 milliliter round bottom flask with a stir bar. Insert the sample-containing thimble into a 40 milliliter soxhlet extractor and add a condenser. Install everything on the parallel extractor work station connected to a recirculating tiller.When all samples are loaded, begin stirring, set the temperature to 135 degrees Celsius and turn on the recirculator. After 20 hours of extraction, turn off the heating to allow refluxing to come to a halt. Then switch both stirring and cooling off.take the thimbles out of the soxhlet using tweazers, and let the wet pulp dry in the thimble overnight in a fume hood. Add the liquid from the soxhlet extraction to the liquid from the biomass wash and continue evaporating the ethanol from the biomass wash with the parallel evaporator or a rotary evaporator at 40 degrees Celsius. After transferring the air-dried pulp from the thimble onto a piece of teared aluminum foil on an analytical balance, record the air-dried weight of the extracted pulp and transfer it into a labeled plastic bag.Try to recover everything while not scraping thimble material off the wall. Determine the moisture content of the pulp immediately to calculate the oven-dried yield as described in the text protocol. When all of the ethanol has evaporated, precipitate the lignin by transferring the ionic liquid from the round bottom flask into a 50 milliliter centrifuge tube using 30 milliliters of water.Mix the suspension and leave it for at least one hour. Then centrifuge the suspension for 20 minutes at 2, 000 times G, and separate the solution from the solid by decanting. Add 30 milliliters of distilled water to the lignin pellet inside the centrifuge tube.Repeat the mixing incubation for one hour and centrifugation. Repeat this step for a total of three lignin washes. Dry the lignin inside the centrifuge tube using a vacuum oven and a pierced lid at 45 degrees Celsius overnight.To determine the lignin yield, place a piece of aluminum foil on the balance, tear the weight, add lignin from the oven, and record the weight immediately. Finally, transfer the lignin into a vial for storage. The amount of pulp and lignin isolated after pretreatment depends on the reaction conditions, the feedstock, and the ionic liquid.A typical time-course is presented here, where pulp yields initially decrease as lignin and hemicelluloses are extracted, while lignin yields increase over time. The lignin is isolated as a brown solid that had a faint but pleasant smell. The properties of the recovered cellulose pulp strongly depend on the conditions chosen.Running the pretreatment for too long or at too high temperatures will result in the degradation of the cellulose and formation of pseudo-lignin. More differences between the pulp properties become evident when subjecting it to further analysis, such as compositional analysis or enzymatic saccharification. Shown here is the compositional analysis of pine pulp and miscanthus pulp pretreated using same tenmerature and time.As can be seen, the pulp composition is very different as a result of the different types of biomass reacting differently to the same conditions. After watching this video, you should have a good understanding of how to perform pretreatment using low-cost ionic liquids and how to study different experimented parameters. Once mastered, this technique can be completed in around five days.This includes a lot of downtime and waiting, so other tasks can be performed in parallel. Following this procedure, other methods, such as compositional analysis of the pulp, saccharification of the pulp, or lignin NMR, and molecular weight analysis can be carried out. These will further characterize the isolated materials.I had the idea for this method when I discovered that certain ionic liquids move lignin from lignocellulosic biomass after absorbing moisture from air. By varying the water content before and after the pretreatment, I found that I could isolate the lignin easily by adding water after the extraction. Don't forget that working with acids and elevated temperatures can be hazardous, and personal protective equipment, such as lab specs, lab coats, and appropriate gloves, should always be worn while performing this procedure.

pretreatment-of-lignocellulosic-biomass-with-low-cost-ionic-liquids

Research

JoVE Journal

Environment

High-throughput Screening of Recalcitrance Variations in Lignocellulosic Biomass: Total Lignin, Lignin Monomers, and Enzymatic Sugar Release

The conversion of lignocellulosic biomass to fuels, chemicals, and other commodities has been explored as one possible pathway toward reductions in the use of non-renewable energy sources. In order to identify which plants, out of a diverse pool, have the desired chemical traits for downstream applications, attributes, such as cellulose and lignin content, or monomeric sugar release following an enzymatic saccharification, must be compared. The experimental and data analysis protocols of the standard methods of analysis can be time-consuming, thereby limiting the number of samples that can be measured. High-throughput (HTP) methods alleviate the shortcomings of the standard methods, and permit the rapid screening of available samples to isolate those possessing the desired traits. This study illustrates the HTP sugar release and pyrolysis-molecular beam mass spectrometry pipelines employed at the National Renewable Energy Lab. These pipelines have enabled the efficient assessment of thousands of plants while decreasing experimental time and costs through reductions in labor and consumables.

Plant cell wall structure and chemistry traits are evaluated to identify ideal feedstocks for biofuels and bio-materials. Standard methods have limitations when applied to large data sets. These high-throughput pretreatment, enzyme saccharification, and pyrolysis-molecular beam mass spectrometry methods compare large numbers of biomass samples with decreased experimental time and cost.

The conversion of lignocellulosic biomass to fuels, chemicals, and other commodities has been explored as one possible pathway toward reductions in the ...

Experimental Protocol for Biodiesel Production with Isolation of Alkenones as Coproducts from Commercial Isochrysis Algal Biomass

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. Biomass-derived biofuels have gained considerable attention in this regard, however first generation biofuels from edible crops like corn ethanol or soybean biodiesel have generally fallen out of favor. There is thus great interest in the development of methods for the production of liquid fuels from domestic and superior non-edible sources. Here we describe a detailed procedure for the production of a purified biodiesel from the marine microalgae Isochrysis. Additionally, a unique suite of lipids known as polyunsaturated long-chain alkenones are isolated in parallel as potentially valuable coproducts to offset the cost of biodiesel production. Multi-kilogram quantities of Isochrysis are purchased from two commercial sources, one as a wet paste (80% water) that is first dried prior to processing, and the other a dry milled powder (95% dry). Lipids are extracted with hexanes in a Soxhlet apparatus to produce an algal oil (&#34;hexane algal oil&#34;) containing both traditional fats (i.e., triglycerides, 46-60% w/w) and alkenones (16-25% w/w). Saponification of the triglycerides in the algal oil allows for separation of the resulting free fatty acids (FFAs) from alkenone-containing neutral lipids. FFAs are then converted to biodiesel (i.e., fatty acid methyl esters, FAMEs) by acid-catalyzed esterification while alkenones are isolated and purified from the neutral lipids by crystallization. We demonstrate that biodiesel from both commercial Isochrysis biomasses have similar but not identical FAME profiles, characterized by elevated polyunsaturated fatty acid contents (approximately 40% w/w). Yields of biodiesel were consistently higher when starting from the Isochrysis wet paste (12% w/w vs. 7% w/w), which can be traced to lower amounts of hexane algal oil obtained from the powdered Isochrysis product.

Experimental Protocol for Biodiesel Production with Isolation of Alkenones as Coproducts from Commercial Isochrysis Algal Biomass

Detailed methods are presented for the production of biodiesel along with the co-isolation of alkenones as valuable coproducts from commercial Isochrysis microalgae.

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. ...

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus (Murayama), Using a Low-Cost, Small, and Easily Constructed Flight Mill

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). Therefore, knowing the dispersal capacity may help inform trapping/tree removal efforts to prevent this disease more effectively. In this study, we measured the flight velocity and duration and estimated the flight distance of the beetle using a newly developed flight mill. The flight mill is low cost, small, and constructed using commonly available items. Both the flight mill arm and its vertical axis comprise a thin needle. A beetle specimen is glued to one tip of the arm using instant glue. The other tip is thick due to being covered with plastic, thus it facilitates the detection of rotations of the arm. The revolution of the arm is detected by a photo sensor mounted on an infrared LED, and is indicated by a change in the output voltage when the arm passed above the LED. The photo sensor is connected to a personal computer and the output voltage data are stored at a sampling rate of 1 kHz. By conducting experiments using this flight mill, we found that P. quercivorus can fly at least 27 km. Because our flight mill comprises cheap and small ordinary items, many flight mills can be prepared and used simultaneously in a small laboratory space. This enables experimenters to obtain a sufficient amount of data within a short period.

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus Murayama, Using a Low-Cost, Small, and Easily Constructed Flight Mill

We developed a low cost and small flight mill, constructed with commonly available items and easily used in experimentation. Using this apparatus, we measured the flight ability of an ambrosia beetle, Platypus quercivorus.

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). ...

Ammonia Fiber Expansion (AFEX) Pretreatment of Lignocellulosic Biomass

Lignocellulosic materials are plant-derived feedstocks, such as crop residues (e.g., corn stover, rice straw, and sugar cane bagasse) and purpose-grown energy crops (e.g., miscanthus, and switchgrass) that are available in large quantities to produce biofuels, biochemicals, and animal feed. Plant polysaccharides (i.e., cellulose, hemicellulose, and pectin) embedded within cell walls are highly recalcitrant towards conversion into useful products. Ammonia fiber expansion (AFEX) is a thermochemical pretreatment that increases accessibility of polysaccharides to enzymes for hydrolysis into fermentable sugars. These released sugars can be converted into fuels and chemicals in a biorefinery. Here, we describe a laboratory-scale batch AFEX process to produce pretreated biomass on the gram-scale without any ammonia recycling. The laboratory-scale process can be used to identify optimal pretreatment conditions (e.g., ammonia loading, water loading, biomass loading, temperature, pressure, residence time, etc.) and generates sufficient quantities of pretreated samples for detailed physicochemical characterization and enzymatic/microbial analysis. The yield of fermentable sugars from enzymatic hydrolysis of corn stover pretreated using the laboratory-scale AFEX process is comparable to pilot-scale AFEX process under similar pretreatment conditions. This paper is intended to provide a detailed standard operating procedure for the safe and consistent operation of laboratory-scale reactors for performing AFEX pretreatment of lignocellulosic biomass.

Ammonia Fiber Expansion AFEX Pretreatment of Lignocellulosic Biomass

Ammonia fiber expansion (AFEX) is a thermochemical pretreatment technology that can convert lignocellulosic biomass (e.g., corn stover, rice straw, and sugarcane bagasse) into a highly digestible feedstock for both biofuels and animal feed applications. Here, we describe a laboratory-scale method for conducting AFEX pretreatment on lignocellulosic biomass.

Lignocellulosic materials are plant-derived feedstocks, such as crop residues (e.g., corn stover, rice straw, and sugar cane bagasse) and purpose-grown ...

A Novel Method for the Pentosan Analysis Present in Jute Biomass and Its Conversion into Sugar Monomers Using Acidic Ionic Liquid

Recently, ionic liquids (ILs) are used for biomass valorization into valuable chemicals because of their remarkable properties such as thermal stability, lower vapor pressure, non-flammability, higher heat capacity, and tunable solubility and acidity. Here, we demonstrate a method for the synthesis of C5 sugars (xylose and arabinose) from the pentosan present in jute biomass in a one-pot process by utilizing a catalytic amount of Br&#248;nsted acidic 1-methyl-3-(3-sulfopropyl)-imidazolium hydrogen sulfate IL. The acidic IL is synthesized in the lab and characterized using NMR spectroscopic techniques for understanding its purity. The various properties of BAIL are measured such as acid strength, thermal and hydrothermal stability, which showed that the catalyst is stable at a higher temperature (250 &#176;C) and possesses very high acid strength (Ho 1.57). The acidic IL converts over 90% of pentosan into sugars and furfural. Hence, the presenting method in this study can also be employed for the evaluation of pentosan concentration in other kinds of lignocellulosic biomass.

We present a protocol for the synthesis of C5 sugars (xylose and arabinose) from a renewable non-edible lignocellulosic biomass (i.e., jute) with the presence of Br&#248;nsted acidic ionic liquids (BAILs) as the catalyst in water. The BAILs catalyst exhibited better catalytic performance than conventional mineral acid catalysts (H2SO4 and HCl).

Recently, ionic liquids (ILs) are used for biomass valorization into valuable chemicals because of their remarkable properties such as thermal stability, ...

A Flexible Low Cost Hydroponic System for Assessing Plant Responses to Small Molecules in Sterile Conditions

A wide range of studies in plant biology are performed using hydroponic cultures. In this work, an in vitro hydroponic growth system designed for assessing plant responses to chemicals and other substances of interest is presented. This system is highly efficient in obtaining homogeneous and healthy seedlings of the C3 and C4 model species Arabidopsis thaliana and Setaria viridis, respectively. The sterile cultivation avoids algae and microorganism contamination, which are known limiting factors for plant normal growth and development in hydroponics. In addition, this system is scalable, enabling the harvest of plant material on a large scale with minor mechanical damage, as well as the harvest of individual parts of a plant if desired. A detailed protocol demonstrating that this system has an easy and low-cost assembly, as it uses pipette racks as the main platform for growing plants, is provided. The feasibility of this system was validated using Arabidopsis seedlings to assess the effect of the drug AZD-8055, a chemical inhibitor of the target of rapamycin (TOR) kinase. TOR inhibition was efficiently detected as early as 30 min after an AZD-8055 treatment in roots and shoots. Furthermore, AZD-8055-treated plants displayed the expected starch-excess phenotype. We proposed this hydroponic system as an ideal method for plant researchers aiming to monitor the action of plant inducers or inhibitors, as well as to assess metabolic fluxes using isotope-labeling compounds which, in general, requires the use of expensive reagents.

A simple, versatile, and low-cost in vitro hydroponic system was successfully optimized, enabling large-scale experiments under sterile conditions. This system facilitates the application of chemicals in a solution and their efficient absorption by roots for molecular, biochemical, and physiological studies.

A wide range of studies in plant biology are performed using hydroponic cultures. In this work, an in vitro hydroponic growth system designed for ...

Preparation of Biomass-based Mesoporous Carbon with Higher Nitrogen-/Oxygen-chelating Adsorption for Cu(II) Through Microwave Pre-Pyrolysis

An environment-friendly technique for synthesizing biomass-based mesoporous activated carbon with high nitrogen-/oxygen-chelating adsorption for Cu(II) is proposed. Bagasse impregnated with phosphoric acid is utilized as the precursor. To pyrolyze the precursor, two separate heating modes are used: microwave pyrolysis and conventional electric-heating pyrolysis. The resulting bagasse-derived carbon samples are modified with nitrification and reduction modification. Nitrogen (N)/oxygen (O) functional groups are simultaneously introduced to the surface of activated carbon, enhancing its adsorption of Cu(II) by complexing and ion-exchange. Characterization and copper adsorption experiments are performed to investigate the physicochemical properties of four prepared carbon samples and determine which heating method favors the subsequent modification for doping of N/O functional groups. In this technique, based on analyzing data of nitrogen adsorption, Fourier transform infrared spectroscopy, and batch adsorption experiments, it is proven that microwave-pyrolyzed carbon has more defect sites and, therefore, time-saving effective microwave pyrolysis contributes more N/O species to the carbon, although it leads to a lower specific surface area. This technique offers a promising route to synthesis adsorbents with higher nitrogen and oxygen content and a higher adsorption capacity of heavy-metal ions in wastewater remediation applications.

Preparation of Biomass-based Mesoporous Carbon with Higher Nitrogen-/Oxygen-chelating Adsorption for CuII Through Microwave Pre-Pyrolysis

Here, we present a protocol to synthesize nitrogen/oxygen dual-doped mesoporous carbon from biomass by chemical activation in different pyrolysis modes followed by modification. We demonstrate that the microwave pyrolysis benefits the subsequent modification process to simultaneously introduce more nitrogen and oxygen functional groups on the carbon.

An environment-friendly technique for synthesizing biomass-based mesoporous activated carbon with high nitrogen-/oxygen-chelating adsorption for Cu(II) is ...

Construction of a Compact Low-Cost Radiation Shield for Air-Temperature Sensors in Ecological Field Studies

Low cost temperature sensors are increasingly used by ecologists to assess climatic variation and change on ecologically relevant scales. Although cost-effective, if not deployed with proper solar radiation shielding, the observations recorded from these sensors will be biased and inaccurate. Manufactured radiation shields are effective at minimizing this bias, but are expensive compared to the cost of these sensors. Here, we provide a detailed methodology for constructing a compact version of a previously described custom fabricated radiation shield, which is more accurate than other published shielding methods that attempt to minimize shield size or construction costs. The method requires very little material: corrugated plastic sheets, aluminum foil duct tape, and cable ties. One 15 cm and two 10 cm squares of corrugated plastic are used for each shield. After cutting, scoring, taping and stapling of the sheets, the 10 cm squares form the bottom two layers of the solar radiation shield, while the 15 cm square forms the top layer. The three sheets are held together with cable ties. This compact solar radiation shield can be suspended, or placed against any flat surface. Care must be taken to ensure that the shield is completely parallel to the ground to prevent direct solar radiation from reaching the sensor, possibly causing increased warm biases in sun-exposed sites in the morning and afternoon relative to the original, larger design. Even so, differences in recorded temperatures between the smaller, compact shield design and the original design were small (mean daytime bias = 0.06 &#176;C). Construction costs are less than half of the original shield design, and the new design results in a less conspicuous instrument that may be advantageous in many field ecology settings.

With the advent of small, low-cost environmental sensors, it is now possible to deploy high-density networks of sensors to measure hyper localized temperature variation. Here, we provide a detailed methodology for constructing a compact version of a previously described custom-fabricated radiation shield for use with inexpensive thermochrons.

Low cost temperature sensors are increasingly used by ecologists to assess climatic variation and change on ecologically relevant scales. Although ...

Construction of a Low-cost Mobile Incubator for Field and Laboratory Use

Incubators are essential for a range of culture-based microbial methods, such as membrane filtration followed by cultivation for assessing drinking water quality. However, commercially available incubators are often costly, difficult to transport, not flexible in terms of volume, and/or poorly adapted to local field conditions where access to electricity is unreliable. The purpose of this study was to develop an adaptable, low-cost and transportable incubator that can be constructed using readily available components. The electronic core of the incubator was first developed. These components were then tested under a range of ambient temperature conditions (3.5 &#176;C - 39 &#176;C) using three types of incubator shells (polystyrene foam box, hard cooler box, and cardboard box covered with a survival blanket). The electronic core showed comparable performance to a standard laboratory incubator in terms of the time required to reach the set temperature, inner temperature stability and spatial dispersion, power consumption, and microbial growth. The incubator set-ups were also effective at moderate and low ambient temperatures (between 3.5 &#176;C and 27 &#176;C), and at high temperatures (39 &#176;C) when the incubator set temperature was higher. This incubator prototype is low-cost (&lt; 300 USD) and adaptable to a variety of materials and volumes. Its demountable structure makes it easy to transport. It can be used in both established laboratories with grid power or in remote settings powered by solar energy or a car battery. It is particularly useful as an equipment option for field laboratories in areas with limited access to resources for water quality monitoring.

This paper describes a method for building an adaptable, low-cost and transportable incubator for microbial testing of drinking water. Our design is based on widely available materials and can operate under a range of field conditions, while still offering the advantages of higher-end laboratory-based models.

Incubators are essential for a range of culture-based microbial methods, such as membrane filtration followed by cultivation for assessing drinking water ...

BtM, a Low-cost Open-source Datalogger to Estimate the Water Content of Nonvascular Cryptogams

Communities of nonvascular cryptogams, such as mosses or lichens, are an important part of the Earth's biodiversity, contributing to the regulation of the carbon and nitrogen cycles in many ecosystems. Being poikilohydric organisms, they do not actively control their internal water content and need a humid environment to activate their metabolism. Therefore, studying water relationships of nonvascular cryptogams is crucial to understand both their diversity patterns and their functions in the ecosystems. We present the BtM datalogger, a low-cost open-source platform for the study of the water content of nonvascular cryptogams. The datalogger is designed to measure ambient temperature, humidity, and conductance from up to eight samples simultaneously. We provide a design for a printed circuit board (PCB), a detailed protocol to assemble the components, and the required source code. All this makes the assembly of the BtM datalogger accessible to any research group, even to those without previous specialized knowledge. Therefore, the design presented here has the potential to help popularize the use of this type of device among ecologists and field biologists.

We present a simple and cost-effective method to build an open-source datalogger that measures the conductance of nonvascular cryptogams together with the environmental temperature and humidity. We describe the hardware design of the datalogger and provide step-by-step assembly instructions, the list of required open-source logging software, the code to run the datalogger, and a calibration protocol.

Communities of nonvascular cryptogams, such as mosses or lichens, are an important part of the Earth's biodiversity, contributing to the regulation of the ...