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Micromechanical properties of biomedical hydrogel for application as microchannel elastomer
Journal of the Mechanical Behavior of Biomedical Materials 77 (2018) 217–224
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Micromechanical properties of biomedical hydrogel for application as microchannel elastomer
MARK
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Ebenezer O. Igea,d, , M. Kiran Rajb, Ademola A. Darec, Suman Chakrabortyb,d a
Department of Mechanical and Mechatronics Engineering, Afe Babalola University, Ado-Ekiti 360001, Nigeria Advanced Technology Development Centre, Indian Institute of Technology, Kharagpur 721302, India c Department of Mechanical Engineering University of Ibadan, Ibadan, Nigeria d Department of Mechanical Engineering, Indian Institute of Technology, Kharagpur 721302, India b
A R T I C L E I N F O
A B S T R A C T
Keywords: Surface roughness Loss modulus Storage modulus Wettability
Polymers are believed to be the building blocks for the creation of the next generation of materials and devices in practically all areas of biomedical research. There are a number of polymers that are being employed in varied applications in microfluidic platform due to the tremendous possibilities for soft matter based elastomers especially in biomedical applications. Polymeric hydrogels have been used as building block in micro-confinements and for specified function such as flow control. The need exists to suitably determine the mechanical characteristics of gel-based materials for possible use as a microchannel elastomer. In this investigation, we describe synthesis procedure, morphological, wettability characterization of hydrogel elastomer synthesized by free-radical polymerization crosslinked over varying acrylamide composition for 10% w/v: 25% w/w, 15% w/v: 25% w/w, 20% w/v: 25% w/w and 25% w/v: 25% w/w respectively. Micromechanical properties such as surface morphology, wettability, and micro-rheological behaviour of hydrogel elastomer using standard protocols was undertaken to determine roughness, contact angle, loss modulus and storage modulus over varied cross-linking of the constituent monomers. The impact of these parameters on flow transport and microchannel structural stability is well delineated in this report. We established that polymeric hydrogel could be a candidate for whole microchannel elastomer with suitable application in areas of tissues and biomedical engineering to mimic native biological transport conduits.
1. Introduction Polymers have been extensively employed in microfluidic platforms leveraging on the advancement in micro-electromechanical systems (MEMS) (Liu, 2007). There are a variety of classes of polymers that have been employed as elastomers in several microfluidic-based investigations. Fakunle and Aguilar (2006) reported the use of organic and inorganic materials like ceramics in the forms of co-fired ceramics (CFC), vitro-ceramics and polymers in microfluidics. In the recent times, polymer-based materials have been well patronized for microfluidics applications. Yu and Shi (2015) fabricated 2D and 3D microfluidic paper-based analytical devices (μPADs) by photolithographypatterning microchannels on a parafilm and subsequently embossing them to paper. Poly dimethyl siloxane (PDMS) is a household name amongst polymers employed in microfluidics owing to the ease of synthesis from the pre-polymer solution made by proportionate cross-linking of sylgard 184 and curing agent. The reason for increasing patronage and
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popularity of PDMS as the material of choice for microfluidic devices is due to its low cost, ease of fabrication, oxygen permeability and optical transparency (Markov et al., 2015). PDMS possesses inherent mechanical characteristics such as low mechanical properties with Young's modulus of 1.32–2.97 MPa, ultimate tensile strength of 3.51–7.65 MPa, compressive modulus of 117.8–186.9 MPa and ultimate compressive strength of 28.4–51.7 GPa. Johnson et al. (2014). PDMS has affinity for small hydrophobic molecules and thus could lead to biomolecule absorption/adsorption from the medium, thus biasing the experimental condition. The permeability of PDMS to water vapor can also lead to media drying and thus change its osmolarity. These issues have limited the application in a number of biomedical-based investigations. To this end, there have been reports on several alternate elastomers which could be near-native biological tissues to enhance reliability investigation in microfluidic-based platforms. Hydrogels have been used in microfluidic platforms since its mention as flow control device in microfluidic channel. For instance, (Eddington and Beebe, 2004) as well as (Casson and Lutolf, 2014)
Correspondence to: College of Engineering, Afe Babalola University, PMB 5454, Ado-Ekiti 360001, Nigeria. E-mail addresses: [email protected], [email protected] (E.O. Ige).
http://dx.doi.org/10.1016/j.jmbbm.2017.09.011 Received 4 July 2017; Received in revised form 30 August 2017; Accepted 6 September 2017 Available online 14 September 2017 1751-6161/ © 2017 Elsevier Ltd. All rights reserved.
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roughness in the range (Colebrook, 1939). Kandlikar (2005b) reassessed the experimental findings of Nikuradse's with special interest on roughness at micrometer scale. Hence, these studies underscored the importance of conduit roughness in flow transport. There are several techniques employed for measurement of surface roughness such as optical or scanning electron microscopy, profilometer, digital holographic microscopy (Moody, 1944) and Atomic Force Microscopy (AFM). In most reports, AFM is preferred over profilometer because the application of the latter results to localized damage to the surface being examined (Montifort et al., 2006). The impact of surface roughness on flow conduits becomes critical with reduction in channel dimension (Sundararajan et al., 2005; Young et al., 2009). At micron-seized length scale, surface roughness is characterized by short wavelengths presented in both amplitude parameters and spatial parameters which forms the basic parameter illustrated in AFM-based analysis. In addition to surface roughness, micro-rheological information of the flow confinement such as storage and loss modulus is being investigated in this work. This is to provide insight to the effect of monomer cross-linking on the compliant (flexible) nature of elastomers used as flow conduits. The wettability effect of polymerization processes on elastomer surfaces with variation in monomer crosslink is presented in this report. In this study, attention is directed to acrylamide-based hydrogel elastomer and characterization procedures of micromechanical properties of polymeric hydrogel elastomer are delineated for the purpose of fluid transport applications in micro-confinements. This investigation contains presents detailed description of standard protocols for parameters like micro-rheology, surface roughness and wettability that could impact on microscale flow transport. The properties of flow conduit such as surface roughness, contact angle and micro-rheology is important for consideration of elastomer especially in a microfluidic platform. These properties provide the information that could predict possibility of resistance to flow transport on elastomer with specified acrylamide composition which is the proposed benefit of the present investigation.
employed hydrogel as a building material for precise biomolecule delivery in microfluidic set-up. Cheng et al. (2007) used hydrogel for biological experiments because of its inherent ability to respond to chemical stimuli. Hydrogels are a class of polymer that could be derived from synthetic and natural materials. Synthetic materials include poly (ethylene oxide) (PEO), poly (vinyl alcohol) (PVA), poly (acrylic acid) (PAA), poly (propylene furmarate-co-ethylene glycol) (P(PF-co-EG)), and polypeptides. Naturally derived gel-based polymers are agarose, alginate, chitosan, collagen, fibrin, gelatin, and hyaluronic acid (HA) (Drury and Mooney, 2003). Poly(acrylamide-co-acrylic acid) hydrogels are relatively easy to produce, more so, reports have mentioned that it could represent a useful matrix for analytical and synthetic surrogate for biological tissues (Faraji et al., 2011). There are a number of tissue engineering applications that have favoured the use of hydrogel because of its biocompatibility. Such applications include tissue engineering matrices, wound dressing, skin recovery and drug transport etc (Elabadawy and Xin, 2017; Caccavo et al., 2015). The intrinsic property of hydrogel such as its modulus of elasticity or stiffness which has been reported as a characteristic of its extracellular matrix alludes to its applications as cell-dependent anchorage (Pompe et al., 2009). Hydrogel has been extensively utilized as extracellular matrix fiber for micro-architecture adventitia of large blood vessels in a richly vacularized micro-environment (González-Díaz and Varghese, 2016). However, polymeric hydrogel is yet to be fully employed as whole elastomer for microchannel despite its excellent biocompatibility criteria, ease of synthesis and cost benefit among others owing to the challenge of fabricating microchannel-like hydrogel structures (Hammer et al., 2013). In the parlance of tissues engineering, vascular constructs are regarded as microchannels. Hence, techniques for vascularization of biocompatible structures such as hydrogel remain a subject of interest because of the prospects of fluid-carrying vessels in applications of biomedical engineering (Bae et al., 2012; Bertassoni et al., 2014). Several authors have employed strategies for fabricating gel-based conduits. You et al. (2016) fabricated cell-laden hydrogel based three dimensional constructs using 3D bio-printing technology. Yang et al. (2016) attempted to fabricate multi-layer vascular chip by utilizing 3d technology to cast hollow L-shaped microchannel constructs for use as vascular conduits which was incorporated with native umbilical vein endothetlial cells. Zhang et al. (2015) addressed the challenge of fabricating synthetic vascular fluidic conduits by employing projectionbased stereolithography to construct 3D biocompatible hydrogel. Du et al. (2011) applied single and cost-effective layer-by-layer sequential technique to assembly hydrogel constructs with an embedded microchannel using photo-lithography processes. Lee et al. (2016) employed sacrificial template to fabricate 3D microvascular channel in a cellpopulated environment using Poly(N-isopropylacrylamide) based hydrogel based on its credible thermoresponsive attribute. Surface roughness as a determinant of flow in microchannels have been well investigated (Fanning, 1877; Fercana et al., 2017). Because of the practical importance of surface roughness in experimental procedure, several techniques have been proposed to measure this property. Kandlikar et al. (2003) employed optical method based on reflected beam intensity profile of the He-Ne laser beam and a fiber optic probe for detection. Rapid optical system based on eximer laser and beam profiler for nano-scale poly-Si thin film surface roughness measurement was introduced (Kandlikar et al., 2005a). The result was obtained with error to be less than 2.1% and the measurement time was shortened by up to 83%. The correlation between surface roughness and fluid flow was first illustrated by the submission of Darcy in the nineteen century that pressure drop and surface roughness are important factor in fluid flow (Ren et al., 2011). In a latter report, Fanning re-affirmed the same conclusion on surface roughness and pressure drop in pipe (Kandlikar and Schmitt, 2005). There are several reports on this subject such as (Jaeger et al., 2012; Nikuradse, 1933) that corroborated the significance of friction factor and relative roughness. The famous Moody's chart was based Colebrook equation and Nikurdase work with relative
2. Materials and method 2.1. Synthesis of polymeric hydrogel There are three stages of the free-radical polymerization procedure employed in this study. The first stage is the synthesis of linear polymer with functional side group; the base monomer reacts with the carboxylic functional group in acrylic acid to form a linear polymer with the carboxylic group as side attachment. In the second stage, crosslinking reaction takes place where a crosslinker agent which is methylene-bis-acrylamide (BIS) reacts with the functional group attached to the side of the linear polymer in the previous stage. A well crosslinked polyacrylamide-based hydrogel polymer matrix is formed when initiators and accelerators are added. At the third stage un-reacted compound and solvent in the network are removed. The equation of chemical reaction of the monomers and the schematics of the polymerization process is highlighted in Fig. 1. Using the above concept, hydrogels were produced using AAc and BIS as based monomers with APS and TEMED as initiators. Hydrogel compositions used were 10% w/v: 25% w/w, 15% w/v: 25% w/w, 20% w/v: 25% w/w and 25% w/v: 25% w/w. These cross-linker ratios when denoted as the percentage weight of monomers (AAm and BIS) to deionized water (DI water) is written as % w/v and when denoted as percentage of solute monomer to weight of AAc present in hydrogel matrix is written as % w/w. These cross-linker ratios are in reference to volume of deionized water and weight of AAc present in the hydrogel matrix. The initiators employed in acrylamide-co-acrylic acid hydrogel were ammonium persulfate (APS) and Tetraethyl dimethyldiamine (TEMED). The concentrations of these reagents used for the entire synthesis of hydrogel are 0.02 nM and 0.03 mM of APS and TEMED respectively. The concentration of APS used was prepared by diluting 218
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2.3. Analysis of hydrogel elastomer The synthesized hydrogel was characterized by determining the visco-elasticity, surface roughness, hydrophilicity behaviour of hydrogel material using well established procedures. The protocols undertaken during each of this analysis are hereby presented. Micro-rheological characterization of the hydrogel elastomer was carried out using micro-rheometer for measurement of selected micromechanical properties such as storage and bulk modulus as a measure of its visco-elasticity. To understand the effect of liquid-like constituent of the hydrogel composition on the surface morphology of the hydrogel, strips surface mapping using direct contact mode of the cantilever tip was carried out. Likewise to understand the effect of acrylic acid composition in the polyelectrolyte elastomer, surface force topology was analyzed using spectrometry mode of the Atomic Force Microscopy (AFM). To study the hydrophilicity/hydrophobicity tendency of the hydrogel microchannel for the synthesis cross-linking ratios considered in this report, contact angle measurement using sessile techniques on a Goniometer was carried out.
Fig. 1. Synthesis of co-Polymerization of Poly(acrylamide-co-acrylic acid) Hydrogel (a) Acrylic Acid (AAc); (b) Acrylamide (AAm); (c) methylene-bis-acrylamide (BIS); (d) Poly (acrylamide-co-acrylic acid).
Table 1 Concentration of pre-polymer solutions of polyacrylamide hydrogel. Monomer/Conc.
10% w/v
15% w/v
20% w/v 25% w/v
AAm (g) AAc (g) BIS (g) DI Water (mL) APS (µL) TEMED (µL)
0.76 0.25 0.014 10 100 50
1.25 0.375 0.014 10 100 50
1.5 0.5 0.014 10 100 50
2.4. Rheometry and micro-mechanics of hydrogel polymer 1.75 0.70 0.014 10 100 50
Rheometers are used as choice instruments in micro-rheological analysis and generally in the field of rheology. They are of two types: controlled-stress and controlled-strain. In the present investigation, Modular Compact Rheometer with model MCR-301 was used for rheometry analysis of polymeric hydrogel. Polymeric hydrogel was placed on the peltier plate while the shaft spindle of the motor was operated pneumatically to depress the parallel plate against the peltier plate with the hydrogel sample sandwiched between the two surfaces. Strain sweep and frequency sweep data of the hydrogel samples were obtained that were then used to determine the rheometry behaviour. A detailed procedure is presented below.
anhydrous solute of APS of five milligrams (5 mg) weighed in a falcon tube with 100 mL DI water The mixture was gently agitated using SPINIX vortex machine for about 5 min. APS solution at 0.02 mM was used for entire procedure. TEMED of mass 0.02430 g was weighed and dissolved in 50 mL DI water and gently shaken for 5 mins or agitated using SPNIX vortex machine for about 2 min to obtain TEMED with concentration of 0.03 mM. Summary of synthesis and fabrication procedure of the cross-liner ratios for monomers and initiators is described below and stated in Table 1.
2.5. Procedure for rheology analysis of hydrogel elastomer using MCR-310 Hydrogel samples with crosslinker-ratios of 25% w/v: 10% w/w, 25% w/v: 15% w/w, 25% w/v: 20% w/w and 25% w/v: 25% w/w were prepared following the procedure highlighted in Section 2.1 above. For varying cross-linker ratios as specified in Table 1 hydrogel was cut into strips of dimension 2 × 2 × 1 cm. Hydrogel strips was placed on the stainless holder of the sample vice of the rheometer as shown in Fig. 2. Compressed air at 2 bar was used to initiate the spindle run for about five minute prior to operation of rheometer controller unit; this is to ensure that spindle speed is uniform and steady. The controller unit was used to select specified range of stress load on the sample on the peltier plate. Parallel plate was allowed to depress on the hydrogel sample on the peltier plate until the sample is destroyed (this is a destructive material test). The stress values are captured as well as properties such
2.2. Fabrication procedure of polymeric hydrogel elastomer Throughout the entire formulation, the concentration of APS and TEMED were kept constant while the constituent monomer was varied to investigate the effect of acrylamide on surface roughness and other transport properties. Synthesis protocols are itemized as follows: (a) Calculation for gel strength as it is required was undertaken to ensure material economy; (b) AAm and BIS were solute weighed as contained in Table 1 below; (c) AAm and BIS were dissolved in 4.5 mL of D.I water; (d)The mixture in (c) above was gently shaken until solution is formed; (e) 25%w/w of acrylic acid (AAc) was added to (d) above with the aid of a micro-pipette of 200 µL size; (f)The mixture in (e) above was shaken gently to avoid/reduce growth of bubble; (g)100 µL of APS was added to (f) with 10 µL of TEMED added likewise before a gentle shake using a subtractive technique which is similar to that reported in Hassan et al. (2014). The gently shaken pre-polymer solution is transferred to the cylindrical mold of outside diameter 500 µm (LP needle, Comet, India) to fabricate channels in the glass-PMDS bonded housing and placed in a temperature-controlled oven for curing at sustained thermal exposure of 60 ± 5◈C for about 30 ± 5 min. This procedure was repeated for other crosslinker ratios considered in this study as listed in Table 1 to show the description of the pre-network formulation conditions of polymeric hydrogen. The magnitude of concentrations of pre-polymer solutions and notations of monomers (w/v and w/w) used in this investigation is similar to what was obtained in literatures (Chanda and Patel, 2011; Orakdogen and Okay, 2006; Annabi et al., 2017).
Controller/ Computer
Spindle
Pneumatic System Display & Printer
Peltier Plate Hydrogel Sample Sample Vice
Fig. 2. Schematic of experimental set-up for micro-rheological investigation.
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as viscosity, bulk modulus and storage modulus of the hydrogel samples. The sample plots showing the stress behaviours during these tests were obtained using the Anton-Paar software and presented in the result section.
2.6. Surface morphology mapping using AFM agilent 5500 model There were four principal parameters that were employed to quantitatively characterize surface morphology in polymeric hydrogel elastomer which are amplitude or height, spacing, hybrid and functional parameters. The Eqs. (1), (2) and (3) presented below were employed to obtain the values of these parameters. These parameters were characterized using average roughness (Ra) and the root mean square roughness (Rq). In three dimensional (3D) topographic surface profile matrix only the arithmetic average mean, average roughness were used as suggested by Chen et al. (2000), Gadelmawlaa et al. (2002) and Maksumov et al. (2004). Arithmetic Average Mean [Z ]
1 Z (N , M ) = NM
N
Fig. 3. Experimental set-up of wettability investigation L-light source; H-hydrogel elastomer; T-vibration-proof Table; P-program/software; C-camera (high speed); S-syringe/ micro-pipette; D-display unit; Dp-droplet.
M
∑ ∑ Z (x , y) (1)
x=1 y=1
2.9. Experimental procedure for determination of wettability on hydrogel elastomer
Average Roughness [Ra]
Ra (N , M ) =
1 NM
N
M
∑ ∑ Z (x , y) Z (N , M )
Hydrogel sample was attached to glass slide. The goniometer was launched into operation and carefully adjusted to focus the needle. A live video was launched to capture the droplet formation and wetting processes on the hydrogel strips. A 4 µL droplet of liquid (DI water) was allowed to drop on the hydrogel strip in increments of 2 µL. The contact angle of the droplet on the wetted hydrogel surface was then measured. The processes highlighted were repeated for all samples.
(2)
x=1 y=1
Root Mean square [Rq]
⎡ 1 Ra (N , M ) = ⎢ NM ⎣
N
M
∑∑ x=1 y=1
1 2
⎤ [Z (x , y ) − Z (N , M )]2 ⎥ ⎦
(3)
Polymeric hydrogel with abundant hydration content in the matrix tends to be hydrophilic because of the presence of AAm as the base monomer (Nesrinne and Djaamel, 2017), therefore, understanding of the surface asperities is of practical importance (Fu et al., 2014). In this study, effort was made to analyze the surface topology of surface hydrogel strip using the tapping mode of Atomic Force Microscopy (AFM).
3. Results and discussion 3.1. Effect of acrylamide composition on the rheological properties of hydrogel Based on the experimental values, plots of the loss modulus (G′) and storage modulus (G″) against machine frequency for varying composition and cross linker ratios (CR) were obtained and presented in Fig. 4(a)-(d). The rheometery plots for this polymeric hydrogel sample showed strain increases at storage modulus below 2.99 × 103 Pa while loss modulus increases within the range of 7.4 × 10° and 1.484 × 10° Hz for strain sweep analysis. The loss and bulk modulus for hydrogel at crosslinker ratio of 15% w/w: 20% w/v (Fig. 4c) was observed to increase linearly with increase in applied stress load on the hydrogel sample. It could be deduced that hydrogel matrix under this polymer crosslink possesses high strain energy that accommodates high stress load (8.1 ×103 Pa) imposed on it via the peltier plate of the rheometer. However, it also gives an indication that the elastic response reduces with increasing load application as could also be observed with hydrogel crosslinked at 15% w/w: 25% w/v (Fig. 4d). Storage modulus decreases while loss modulus increases sharply at the onset of stress load application which is comparatively lower than that at crosslinker ratio of 15% w/w: 20% w/v (see Fig. 4d). The increase in monomer hydrate load (AAc) at this crosslinked condition could suggest considerably lower strain energy in the hydrogel matrix during the rheometry analysis. Increasing acrylic acid constituent in polymerised hydrogel increases the presence of free (mobile) ions over fixed ligand in hydrogel matrix. The elastic modulus of cross-linker ratios obtained using the relation according to Hosseini et al. (2014a) was in the range of 1.003–1.0047. This indicated that variation of concentration acrylamide monomer effect the structural property of polymeric hydrogel is
2.7. Experimental surface morphology mapping on hydrogel elastomer Hydrogel samples with cross-linker ratios of 25% w/v: 10% w/w, 25% w/v: 15% w/w, 25% w/v: 20% w/w and 25% w/v: 25% w/w were prepared. Hydrogel samples were fastened on 2 × 2 cm glass slide piece using double side adhesion tape. Glass slide-hydrogel sample assembly above was placed gently on the vice of the Agilent 550 Model of AFM and attached securely using the vice brackets. The machine pallet was firmly secured, then the AFM control system was used to launch the machine into operation; Preliminary run of the cantilever tip over the hydrogel sample was undertaken and the run and retract strokes were mapped on each other to check if they are in-phase. Roughness parameters such as mean peak height; mean pit height and root mean square height were thus obtained.
2.8. Wettability analysis on hydrogel elastomer The presence of hydrate load in the polymer constituent suggests hydrophilicity in the hydrogel matrix. This characteristic has the potential to impart on the flow behaviour in hydrogel microchannel. The contact angle measurement can be used to measure the hydrophilicity. Goniometer was used in this research to measure the contact angle. A pictorial and schematic setup of the experiment is shown in Fig. 3 respectively. 220
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Fig. 4. Plots of the loss modulus (G′) and storage modulus (G″) as Functions of Frequency for varying Compositions. (a) 15%w/w: 15%w/v; (b) 15%w/w: 20%w/v; (c) 15%w/w: 20%w/ v; (d) 15%w/w: 25%w/v AAm: AAc Crosslinker Ratio.
candidate elastomer for compliant conduits in investigations relating to tissue and biomedical engineering. 3.2. Surface characteristics of the hydrogel at varying composition A pictorial view of the hydrogel surface for varying composition is shown in Fig. 6. The analysis of AFM results was carried out using Ra, Rq earlier mentioned in section. A summary of this is presented in Table 2. The magnification of objective view of the AFM device which is the interrogation window (IW) set at 90 × 90 µm represented in Fig. 6(a-d) to allow an improved surface mapping yielded root mean square height of 0.0974 µm, maximum peak height of 0.78 µm and maximum pit height of 0.447 µm for hydrogel sample at concentration of 15% w/v-25% w/w. The heights parameters recorded for the 90 × 90 µm as shown in Fig. 6 yielded root mean square height of 0.196 µm, maximum peak height of 0.78 µm and maximum pit height of 0.447 µm. The difference in height parameters for this gel strength was observed to be in the order of one magnitude while the difference in maximum peak heights for the two IW was recorded as mean deviation of 55.6%. At crosslinker ratio of 20% w/v: 25% w/w hydrogel samples with this cross-linker strength at 90 × 90 µm as analyzed. The height parameters were obtained as root mean square height of 17.6 nm; mean peak height at 66.6 nm and mea pit height of 139 nm. For hydrogel with concentration of 25% w/v: 25% w/w, using cantilever tip scan window of 90 × 90 µm, heights parameters recorded were root mean
Fig. 5. Comparative plot showing the rheological behaviour of the hydrogels for two different compositions, S1 denotes for AAc: AAm 10% w/v: 25% w/w whereas S3 denotes 20% w/v: 25% w/w. The measurement was performed in strain sweep mode.
in the scale 10−3. The plot indicated as S1 in Fig. 5 showed that both elastic modulus (G′) and viscous modulus (G″) are independent of shearing time and it represents the stiffness of the hydrogel which is an indication of co-existence of both viscous and elastic properties. While for S3, it was observed that G′ increased over G″ by order one magnitude at low shearing time. But over sustained shearing, both elastic and viscous modulus assumed time independence. These characteristics properties could be beneficial for the consideration of hydrogel as a
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Fig. 6. Characteristics of surface topology measured through atomic force microscopy (AFM) on hy. (a) 15%w/w: 15%w/v; (b) 15%w/w: 20%w/v; (c) 15%w/w: 20%w/v; (d) 15%w/w: 25%w/v AAm: AAc Crosslinker Ratio.
bearing liquid film of the liquid drop causing the spreading of the liquid layer over the hydrogel surface (Hoffman, 2012). This property supports in-vivo cell culture in vascular networks that are hydrophilic in nature (Liu et al., 2013). The results presented in Table 2 showed the decreasing contact angles values, RMS of AFM for varying cross-linker ratio of the gelbased microchannel elastomer. The resulted showed that hydrogel matrix surface roughness decreases with increasing cross-linker ratio implying that hydrophilicity of the matrix increases with increasing concentration of acrylic acid in the polyacrylamide-based hydrogel (Enas, 2015; Farooqi et al., 2017). We observed that wetting property impacted by variation in acrylamide composition influences surface roughness and this could be implicated to the actions of intermolecular forces in the hydrogel matrix (Chakraborty et al., 2012).
Table 2 Comparison of wettability, surface roughness. Hydrogel specification (AAm_AAc ratio)
Contact angle (in degrees)
RMS height (from AFM measurements) µm
10% 15% 20% 25%
66 68 72 74
0.196 0.97 0.17 0.025
w/v_25% w/v_25% w/v_25% w/v_25%
w/w w/w w/w w/w
square height of 25.5 nm, mean peak height 150 nm and mean pit height of 51 nm. This value implies improved surface roughness on the hydrogel with increasing concentration of acrylic acid constituent of the polymeric hydrogel samples analyzed which is in consistent to literature report (see Hosseini et al., 2014b; Farahmand et al., 2015; Boks et al., 2008).
3.4. Comparative flow profiles of velocity distribution in hydrogel and PDMS micro-conduits
3.3. Effect of acrylamide composition of surface wettability A preliminary flow investigation was conducted to observe the behaviour of flow using hydrogel elastomer for selected flow rate. The behaviour of flow field at 1 µL/min for PDMS crosslinker ratio of 30:1 as captured in Fig. 7A showed the flow experienced very strong recirculation between channel axial lengths of 0.13–0.27 cm. An abrupt obstruction encountered by the flow was short-lived while strong recirculation energises the flow downstream the channel. Within the domain of flow, velocity magnitude which was high at the channel entry region suffered early decay probably due to the presence of unpolymerised gel in the flow domain (see Fig. 7B). However, recirculation occurred at about 0.1 unit length of the flow channel and velocity
Although hydrophilicity of the hydrogel surface could be assessed using the RMS technique, a more assuring test by determination of the surface contact angles were carried out as earlier discussed. Table 2 contained the contact angle values for varying acrylamide composition in the hydrogel. It could be observed that contact angle decreases with increasing acrylic acid content in the hydrogel matrix which is the hydrate constituent in the pre-polymer solution. It is also concluded that since contact angles were below 90 degree, that the surface is hydrophilic irrespective of the composition (Cha et al., 2013). There is thus a strong adhesive force between the hydrogel surface and the 222
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Fig. 7. A: Plot of velocity magnitude in PDMS microchannel of CR30:1 at flow rate of 1 µL/min. Fig. 7B: Velocity magnitude at flow rate of Q1 in 10% w/v: 25% w/w hydrogel microchannel at flow rate of 1 µL/min. Fig. 8. Comparative plot of flow velocity distribution in PDMS-hydrogel microchannel at 1 µL/min.
growth was rapid as the flow exit the PDMS-based microchannel. The comparative plots for both PDMS and Hydrogel at considerably near cross linker ratio presented in Fig. 8 showed that fluid flow in both microchannels assumed a similar profile. The maximum velocity attained was an order of magnitude lower in hydrogel microchannel. This could be alluded to the compliant nature of the hydrogel elastomer and could suggest a probable use to investigate flow where importance is attached to the flexible nature wall.
microchannels within the selected range of elastomer composition. Wettability of hydrogel elastomer shifts gradually within the hydrophilicity range with a mean deviation of 2.7 degree. Hydrogel elastomer is here presented as candidate elastomer for whole microchannel fabrication for studies where flexibility specifically fluid-channel flow compliance and hydrophilicity are of critical requirements.
4. Conclusion
This work was supported by the Department of Science and Technology of The Government of India and Federation of Indian Chamber of Commerce and Industries (FICI) through CV Raman International Fellowship awarded to the first author (Ebenezer Olubunmi Ige). The authors are grateful to Indian Institute of Technology Kharagpur for providing the facilities used in this research. Mr Sudipto Panda, Dr Shantimoy Kar and Prof. Sanatanu Chatophadhahy are well appreciated for their contribution to this work.
Acknowledgement
Hydrogel elastomer has been subjected to surface analysis to determine the visco-elastic, surface roughness and wettability behaviour under Newtonian fluid flow condition. The results obtained showed that storage modulus increases with increasing acrylamide composition which suggests improved structural rigidity when such elastomer is used as flow confinement in fluidic system and as replacement or mimic-agents of native biological conduits. Likewise, the visco-elastic response presents hydrogel as candidate elastomer for biomimetic studies of native flexible and compliant channels. The growth in surface roughness for morphology analysis with reducing acrylamide composition indicated the possibility of resistance to fluidic motion in
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Micromechanical properties of biomedical hydrogel for application as microchannel elastomer
MARK
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Ebenezer O. Igea,d, , M. Kiran Rajb, Ademola A. Darec, Suman Chakrabortyb,d a
Department of Mechanical and Mechatronics Engineering, Afe Babalola University, Ado-Ekiti 360001, Nigeria Advanced Technology Development Centre, Indian Institute of Technology, Kharagpur 721302, India c Department of Mechanical Engineering University of Ibadan, Ibadan, Nigeria d Department of Mechanical Engineering, Indian Institute of Technology, Kharagpur 721302, India b
A R T I C L E I N F O
A B S T R A C T
Keywords: Surface roughness Loss modulus Storage modulus Wettability
Polymers are believed to be the building blocks for the creation of the next generation of materials and devices in practically all areas of biomedical research. There are a number of polymers that are being employed in varied applications in microfluidic platform due to the tremendous possibilities for soft matter based elastomers especially in biomedical applications. Polymeric hydrogels have been used as building block in micro-confinements and for specified function such as flow control. The need exists to suitably determine the mechanical characteristics of gel-based materials for possible use as a microchannel elastomer. In this investigation, we describe synthesis procedure, morphological, wettability characterization of hydrogel elastomer synthesized by free-radical polymerization crosslinked over varying acrylamide composition for 10% w/v: 25% w/w, 15% w/v: 25% w/w, 20% w/v: 25% w/w and 25% w/v: 25% w/w respectively. Micromechanical properties such as surface morphology, wettability, and micro-rheological behaviour of hydrogel elastomer using standard protocols was undertaken to determine roughness, contact angle, loss modulus and storage modulus over varied cross-linking of the constituent monomers. The impact of these parameters on flow transport and microchannel structural stability is well delineated in this report. We established that polymeric hydrogel could be a candidate for whole microchannel elastomer with suitable application in areas of tissues and biomedical engineering to mimic native biological transport conduits.
1. Introduction Polymers have been extensively employed in microfluidic platforms leveraging on the advancement in micro-electromechanical systems (MEMS) (Liu, 2007). There are a variety of classes of polymers that have been employed as elastomers in several microfluidic-based investigations. Fakunle and Aguilar (2006) reported the use of organic and inorganic materials like ceramics in the forms of co-fired ceramics (CFC), vitro-ceramics and polymers in microfluidics. In the recent times, polymer-based materials have been well patronized for microfluidics applications. Yu and Shi (2015) fabricated 2D and 3D microfluidic paper-based analytical devices (μPADs) by photolithographypatterning microchannels on a parafilm and subsequently embossing them to paper. Poly dimethyl siloxane (PDMS) is a household name amongst polymers employed in microfluidics owing to the ease of synthesis from the pre-polymer solution made by proportionate cross-linking of sylgard 184 and curing agent. The reason for increasing patronage and
⁎
popularity of PDMS as the material of choice for microfluidic devices is due to its low cost, ease of fabrication, oxygen permeability and optical transparency (Markov et al., 2015). PDMS possesses inherent mechanical characteristics such as low mechanical properties with Young's modulus of 1.32–2.97 MPa, ultimate tensile strength of 3.51–7.65 MPa, compressive modulus of 117.8–186.9 MPa and ultimate compressive strength of 28.4–51.7 GPa. Johnson et al. (2014). PDMS has affinity for small hydrophobic molecules and thus could lead to biomolecule absorption/adsorption from the medium, thus biasing the experimental condition. The permeability of PDMS to water vapor can also lead to media drying and thus change its osmolarity. These issues have limited the application in a number of biomedical-based investigations. To this end, there have been reports on several alternate elastomers which could be near-native biological tissues to enhance reliability investigation in microfluidic-based platforms. Hydrogels have been used in microfluidic platforms since its mention as flow control device in microfluidic channel. For instance, (Eddington and Beebe, 2004) as well as (Casson and Lutolf, 2014)
Correspondence to: College of Engineering, Afe Babalola University, PMB 5454, Ado-Ekiti 360001, Nigeria. E-mail addresses: [email protected], [email protected] (E.O. Ige).
http://dx.doi.org/10.1016/j.jmbbm.2017.09.011 Received 4 July 2017; Received in revised form 30 August 2017; Accepted 6 September 2017 Available online 14 September 2017 1751-6161/ © 2017 Elsevier Ltd. All rights reserved.
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roughness in the range (Colebrook, 1939). Kandlikar (2005b) reassessed the experimental findings of Nikuradse's with special interest on roughness at micrometer scale. Hence, these studies underscored the importance of conduit roughness in flow transport. There are several techniques employed for measurement of surface roughness such as optical or scanning electron microscopy, profilometer, digital holographic microscopy (Moody, 1944) and Atomic Force Microscopy (AFM). In most reports, AFM is preferred over profilometer because the application of the latter results to localized damage to the surface being examined (Montifort et al., 2006). The impact of surface roughness on flow conduits becomes critical with reduction in channel dimension (Sundararajan et al., 2005; Young et al., 2009). At micron-seized length scale, surface roughness is characterized by short wavelengths presented in both amplitude parameters and spatial parameters which forms the basic parameter illustrated in AFM-based analysis. In addition to surface roughness, micro-rheological information of the flow confinement such as storage and loss modulus is being investigated in this work. This is to provide insight to the effect of monomer cross-linking on the compliant (flexible) nature of elastomers used as flow conduits. The wettability effect of polymerization processes on elastomer surfaces with variation in monomer crosslink is presented in this report. In this study, attention is directed to acrylamide-based hydrogel elastomer and characterization procedures of micromechanical properties of polymeric hydrogel elastomer are delineated for the purpose of fluid transport applications in micro-confinements. This investigation contains presents detailed description of standard protocols for parameters like micro-rheology, surface roughness and wettability that could impact on microscale flow transport. The properties of flow conduit such as surface roughness, contact angle and micro-rheology is important for consideration of elastomer especially in a microfluidic platform. These properties provide the information that could predict possibility of resistance to flow transport on elastomer with specified acrylamide composition which is the proposed benefit of the present investigation.
employed hydrogel as a building material for precise biomolecule delivery in microfluidic set-up. Cheng et al. (2007) used hydrogel for biological experiments because of its inherent ability to respond to chemical stimuli. Hydrogels are a class of polymer that could be derived from synthetic and natural materials. Synthetic materials include poly (ethylene oxide) (PEO), poly (vinyl alcohol) (PVA), poly (acrylic acid) (PAA), poly (propylene furmarate-co-ethylene glycol) (P(PF-co-EG)), and polypeptides. Naturally derived gel-based polymers are agarose, alginate, chitosan, collagen, fibrin, gelatin, and hyaluronic acid (HA) (Drury and Mooney, 2003). Poly(acrylamide-co-acrylic acid) hydrogels are relatively easy to produce, more so, reports have mentioned that it could represent a useful matrix for analytical and synthetic surrogate for biological tissues (Faraji et al., 2011). There are a number of tissue engineering applications that have favoured the use of hydrogel because of its biocompatibility. Such applications include tissue engineering matrices, wound dressing, skin recovery and drug transport etc (Elabadawy and Xin, 2017; Caccavo et al., 2015). The intrinsic property of hydrogel such as its modulus of elasticity or stiffness which has been reported as a characteristic of its extracellular matrix alludes to its applications as cell-dependent anchorage (Pompe et al., 2009). Hydrogel has been extensively utilized as extracellular matrix fiber for micro-architecture adventitia of large blood vessels in a richly vacularized micro-environment (González-Díaz and Varghese, 2016). However, polymeric hydrogel is yet to be fully employed as whole elastomer for microchannel despite its excellent biocompatibility criteria, ease of synthesis and cost benefit among others owing to the challenge of fabricating microchannel-like hydrogel structures (Hammer et al., 2013). In the parlance of tissues engineering, vascular constructs are regarded as microchannels. Hence, techniques for vascularization of biocompatible structures such as hydrogel remain a subject of interest because of the prospects of fluid-carrying vessels in applications of biomedical engineering (Bae et al., 2012; Bertassoni et al., 2014). Several authors have employed strategies for fabricating gel-based conduits. You et al. (2016) fabricated cell-laden hydrogel based three dimensional constructs using 3D bio-printing technology. Yang et al. (2016) attempted to fabricate multi-layer vascular chip by utilizing 3d technology to cast hollow L-shaped microchannel constructs for use as vascular conduits which was incorporated with native umbilical vein endothetlial cells. Zhang et al. (2015) addressed the challenge of fabricating synthetic vascular fluidic conduits by employing projectionbased stereolithography to construct 3D biocompatible hydrogel. Du et al. (2011) applied single and cost-effective layer-by-layer sequential technique to assembly hydrogel constructs with an embedded microchannel using photo-lithography processes. Lee et al. (2016) employed sacrificial template to fabricate 3D microvascular channel in a cellpopulated environment using Poly(N-isopropylacrylamide) based hydrogel based on its credible thermoresponsive attribute. Surface roughness as a determinant of flow in microchannels have been well investigated (Fanning, 1877; Fercana et al., 2017). Because of the practical importance of surface roughness in experimental procedure, several techniques have been proposed to measure this property. Kandlikar et al. (2003) employed optical method based on reflected beam intensity profile of the He-Ne laser beam and a fiber optic probe for detection. Rapid optical system based on eximer laser and beam profiler for nano-scale poly-Si thin film surface roughness measurement was introduced (Kandlikar et al., 2005a). The result was obtained with error to be less than 2.1% and the measurement time was shortened by up to 83%. The correlation between surface roughness and fluid flow was first illustrated by the submission of Darcy in the nineteen century that pressure drop and surface roughness are important factor in fluid flow (Ren et al., 2011). In a latter report, Fanning re-affirmed the same conclusion on surface roughness and pressure drop in pipe (Kandlikar and Schmitt, 2005). There are several reports on this subject such as (Jaeger et al., 2012; Nikuradse, 1933) that corroborated the significance of friction factor and relative roughness. The famous Moody's chart was based Colebrook equation and Nikurdase work with relative
2. Materials and method 2.1. Synthesis of polymeric hydrogel There are three stages of the free-radical polymerization procedure employed in this study. The first stage is the synthesis of linear polymer with functional side group; the base monomer reacts with the carboxylic functional group in acrylic acid to form a linear polymer with the carboxylic group as side attachment. In the second stage, crosslinking reaction takes place where a crosslinker agent which is methylene-bis-acrylamide (BIS) reacts with the functional group attached to the side of the linear polymer in the previous stage. A well crosslinked polyacrylamide-based hydrogel polymer matrix is formed when initiators and accelerators are added. At the third stage un-reacted compound and solvent in the network are removed. The equation of chemical reaction of the monomers and the schematics of the polymerization process is highlighted in Fig. 1. Using the above concept, hydrogels were produced using AAc and BIS as based monomers with APS and TEMED as initiators. Hydrogel compositions used were 10% w/v: 25% w/w, 15% w/v: 25% w/w, 20% w/v: 25% w/w and 25% w/v: 25% w/w. These cross-linker ratios when denoted as the percentage weight of monomers (AAm and BIS) to deionized water (DI water) is written as % w/v and when denoted as percentage of solute monomer to weight of AAc present in hydrogel matrix is written as % w/w. These cross-linker ratios are in reference to volume of deionized water and weight of AAc present in the hydrogel matrix. The initiators employed in acrylamide-co-acrylic acid hydrogel were ammonium persulfate (APS) and Tetraethyl dimethyldiamine (TEMED). The concentrations of these reagents used for the entire synthesis of hydrogel are 0.02 nM and 0.03 mM of APS and TEMED respectively. The concentration of APS used was prepared by diluting 218
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2.3. Analysis of hydrogel elastomer The synthesized hydrogel was characterized by determining the visco-elasticity, surface roughness, hydrophilicity behaviour of hydrogel material using well established procedures. The protocols undertaken during each of this analysis are hereby presented. Micro-rheological characterization of the hydrogel elastomer was carried out using micro-rheometer for measurement of selected micromechanical properties such as storage and bulk modulus as a measure of its visco-elasticity. To understand the effect of liquid-like constituent of the hydrogel composition on the surface morphology of the hydrogel, strips surface mapping using direct contact mode of the cantilever tip was carried out. Likewise to understand the effect of acrylic acid composition in the polyelectrolyte elastomer, surface force topology was analyzed using spectrometry mode of the Atomic Force Microscopy (AFM). To study the hydrophilicity/hydrophobicity tendency of the hydrogel microchannel for the synthesis cross-linking ratios considered in this report, contact angle measurement using sessile techniques on a Goniometer was carried out.
Fig. 1. Synthesis of co-Polymerization of Poly(acrylamide-co-acrylic acid) Hydrogel (a) Acrylic Acid (AAc); (b) Acrylamide (AAm); (c) methylene-bis-acrylamide (BIS); (d) Poly (acrylamide-co-acrylic acid).
Table 1 Concentration of pre-polymer solutions of polyacrylamide hydrogel. Monomer/Conc.
10% w/v
15% w/v
20% w/v 25% w/v
AAm (g) AAc (g) BIS (g) DI Water (mL) APS (µL) TEMED (µL)
0.76 0.25 0.014 10 100 50
1.25 0.375 0.014 10 100 50
1.5 0.5 0.014 10 100 50
2.4. Rheometry and micro-mechanics of hydrogel polymer 1.75 0.70 0.014 10 100 50
Rheometers are used as choice instruments in micro-rheological analysis and generally in the field of rheology. They are of two types: controlled-stress and controlled-strain. In the present investigation, Modular Compact Rheometer with model MCR-301 was used for rheometry analysis of polymeric hydrogel. Polymeric hydrogel was placed on the peltier plate while the shaft spindle of the motor was operated pneumatically to depress the parallel plate against the peltier plate with the hydrogel sample sandwiched between the two surfaces. Strain sweep and frequency sweep data of the hydrogel samples were obtained that were then used to determine the rheometry behaviour. A detailed procedure is presented below.
anhydrous solute of APS of five milligrams (5 mg) weighed in a falcon tube with 100 mL DI water The mixture was gently agitated using SPINIX vortex machine for about 5 min. APS solution at 0.02 mM was used for entire procedure. TEMED of mass 0.02430 g was weighed and dissolved in 50 mL DI water and gently shaken for 5 mins or agitated using SPNIX vortex machine for about 2 min to obtain TEMED with concentration of 0.03 mM. Summary of synthesis and fabrication procedure of the cross-liner ratios for monomers and initiators is described below and stated in Table 1.
2.5. Procedure for rheology analysis of hydrogel elastomer using MCR-310 Hydrogel samples with crosslinker-ratios of 25% w/v: 10% w/w, 25% w/v: 15% w/w, 25% w/v: 20% w/w and 25% w/v: 25% w/w were prepared following the procedure highlighted in Section 2.1 above. For varying cross-linker ratios as specified in Table 1 hydrogel was cut into strips of dimension 2 × 2 × 1 cm. Hydrogel strips was placed on the stainless holder of the sample vice of the rheometer as shown in Fig. 2. Compressed air at 2 bar was used to initiate the spindle run for about five minute prior to operation of rheometer controller unit; this is to ensure that spindle speed is uniform and steady. The controller unit was used to select specified range of stress load on the sample on the peltier plate. Parallel plate was allowed to depress on the hydrogel sample on the peltier plate until the sample is destroyed (this is a destructive material test). The stress values are captured as well as properties such
2.2. Fabrication procedure of polymeric hydrogel elastomer Throughout the entire formulation, the concentration of APS and TEMED were kept constant while the constituent monomer was varied to investigate the effect of acrylamide on surface roughness and other transport properties. Synthesis protocols are itemized as follows: (a) Calculation for gel strength as it is required was undertaken to ensure material economy; (b) AAm and BIS were solute weighed as contained in Table 1 below; (c) AAm and BIS were dissolved in 4.5 mL of D.I water; (d)The mixture in (c) above was gently shaken until solution is formed; (e) 25%w/w of acrylic acid (AAc) was added to (d) above with the aid of a micro-pipette of 200 µL size; (f)The mixture in (e) above was shaken gently to avoid/reduce growth of bubble; (g)100 µL of APS was added to (f) with 10 µL of TEMED added likewise before a gentle shake using a subtractive technique which is similar to that reported in Hassan et al. (2014). The gently shaken pre-polymer solution is transferred to the cylindrical mold of outside diameter 500 µm (LP needle, Comet, India) to fabricate channels in the glass-PMDS bonded housing and placed in a temperature-controlled oven for curing at sustained thermal exposure of 60 ± 5◈C for about 30 ± 5 min. This procedure was repeated for other crosslinker ratios considered in this study as listed in Table 1 to show the description of the pre-network formulation conditions of polymeric hydrogen. The magnitude of concentrations of pre-polymer solutions and notations of monomers (w/v and w/w) used in this investigation is similar to what was obtained in literatures (Chanda and Patel, 2011; Orakdogen and Okay, 2006; Annabi et al., 2017).
Controller/ Computer
Spindle
Pneumatic System Display & Printer
Peltier Plate Hydrogel Sample Sample Vice
Fig. 2. Schematic of experimental set-up for micro-rheological investigation.
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as viscosity, bulk modulus and storage modulus of the hydrogel samples. The sample plots showing the stress behaviours during these tests were obtained using the Anton-Paar software and presented in the result section.
2.6. Surface morphology mapping using AFM agilent 5500 model There were four principal parameters that were employed to quantitatively characterize surface morphology in polymeric hydrogel elastomer which are amplitude or height, spacing, hybrid and functional parameters. The Eqs. (1), (2) and (3) presented below were employed to obtain the values of these parameters. These parameters were characterized using average roughness (Ra) and the root mean square roughness (Rq). In three dimensional (3D) topographic surface profile matrix only the arithmetic average mean, average roughness were used as suggested by Chen et al. (2000), Gadelmawlaa et al. (2002) and Maksumov et al. (2004). Arithmetic Average Mean [Z ]
1 Z (N , M ) = NM
N
Fig. 3. Experimental set-up of wettability investigation L-light source; H-hydrogel elastomer; T-vibration-proof Table; P-program/software; C-camera (high speed); S-syringe/ micro-pipette; D-display unit; Dp-droplet.
M
∑ ∑ Z (x , y) (1)
x=1 y=1
2.9. Experimental procedure for determination of wettability on hydrogel elastomer
Average Roughness [Ra]
Ra (N , M ) =
1 NM
N
M
∑ ∑ Z (x , y) Z (N , M )
Hydrogel sample was attached to glass slide. The goniometer was launched into operation and carefully adjusted to focus the needle. A live video was launched to capture the droplet formation and wetting processes on the hydrogel strips. A 4 µL droplet of liquid (DI water) was allowed to drop on the hydrogel strip in increments of 2 µL. The contact angle of the droplet on the wetted hydrogel surface was then measured. The processes highlighted were repeated for all samples.
(2)
x=1 y=1
Root Mean square [Rq]
⎡ 1 Ra (N , M ) = ⎢ NM ⎣
N
M
∑∑ x=1 y=1
1 2
⎤ [Z (x , y ) − Z (N , M )]2 ⎥ ⎦
(3)
Polymeric hydrogel with abundant hydration content in the matrix tends to be hydrophilic because of the presence of AAm as the base monomer (Nesrinne and Djaamel, 2017), therefore, understanding of the surface asperities is of practical importance (Fu et al., 2014). In this study, effort was made to analyze the surface topology of surface hydrogel strip using the tapping mode of Atomic Force Microscopy (AFM).
3. Results and discussion 3.1. Effect of acrylamide composition on the rheological properties of hydrogel Based on the experimental values, plots of the loss modulus (G′) and storage modulus (G″) against machine frequency for varying composition and cross linker ratios (CR) were obtained and presented in Fig. 4(a)-(d). The rheometery plots for this polymeric hydrogel sample showed strain increases at storage modulus below 2.99 × 103 Pa while loss modulus increases within the range of 7.4 × 10° and 1.484 × 10° Hz for strain sweep analysis. The loss and bulk modulus for hydrogel at crosslinker ratio of 15% w/w: 20% w/v (Fig. 4c) was observed to increase linearly with increase in applied stress load on the hydrogel sample. It could be deduced that hydrogel matrix under this polymer crosslink possesses high strain energy that accommodates high stress load (8.1 ×103 Pa) imposed on it via the peltier plate of the rheometer. However, it also gives an indication that the elastic response reduces with increasing load application as could also be observed with hydrogel crosslinked at 15% w/w: 25% w/v (Fig. 4d). Storage modulus decreases while loss modulus increases sharply at the onset of stress load application which is comparatively lower than that at crosslinker ratio of 15% w/w: 20% w/v (see Fig. 4d). The increase in monomer hydrate load (AAc) at this crosslinked condition could suggest considerably lower strain energy in the hydrogel matrix during the rheometry analysis. Increasing acrylic acid constituent in polymerised hydrogel increases the presence of free (mobile) ions over fixed ligand in hydrogel matrix. The elastic modulus of cross-linker ratios obtained using the relation according to Hosseini et al. (2014a) was in the range of 1.003–1.0047. This indicated that variation of concentration acrylamide monomer effect the structural property of polymeric hydrogel is
2.7. Experimental surface morphology mapping on hydrogel elastomer Hydrogel samples with cross-linker ratios of 25% w/v: 10% w/w, 25% w/v: 15% w/w, 25% w/v: 20% w/w and 25% w/v: 25% w/w were prepared. Hydrogel samples were fastened on 2 × 2 cm glass slide piece using double side adhesion tape. Glass slide-hydrogel sample assembly above was placed gently on the vice of the Agilent 550 Model of AFM and attached securely using the vice brackets. The machine pallet was firmly secured, then the AFM control system was used to launch the machine into operation; Preliminary run of the cantilever tip over the hydrogel sample was undertaken and the run and retract strokes were mapped on each other to check if they are in-phase. Roughness parameters such as mean peak height; mean pit height and root mean square height were thus obtained.
2.8. Wettability analysis on hydrogel elastomer The presence of hydrate load in the polymer constituent suggests hydrophilicity in the hydrogel matrix. This characteristic has the potential to impart on the flow behaviour in hydrogel microchannel. The contact angle measurement can be used to measure the hydrophilicity. Goniometer was used in this research to measure the contact angle. A pictorial and schematic setup of the experiment is shown in Fig. 3 respectively. 220
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Fig. 4. Plots of the loss modulus (G′) and storage modulus (G″) as Functions of Frequency for varying Compositions. (a) 15%w/w: 15%w/v; (b) 15%w/w: 20%w/v; (c) 15%w/w: 20%w/ v; (d) 15%w/w: 25%w/v AAm: AAc Crosslinker Ratio.
candidate elastomer for compliant conduits in investigations relating to tissue and biomedical engineering. 3.2. Surface characteristics of the hydrogel at varying composition A pictorial view of the hydrogel surface for varying composition is shown in Fig. 6. The analysis of AFM results was carried out using Ra, Rq earlier mentioned in section. A summary of this is presented in Table 2. The magnification of objective view of the AFM device which is the interrogation window (IW) set at 90 × 90 µm represented in Fig. 6(a-d) to allow an improved surface mapping yielded root mean square height of 0.0974 µm, maximum peak height of 0.78 µm and maximum pit height of 0.447 µm for hydrogel sample at concentration of 15% w/v-25% w/w. The heights parameters recorded for the 90 × 90 µm as shown in Fig. 6 yielded root mean square height of 0.196 µm, maximum peak height of 0.78 µm and maximum pit height of 0.447 µm. The difference in height parameters for this gel strength was observed to be in the order of one magnitude while the difference in maximum peak heights for the two IW was recorded as mean deviation of 55.6%. At crosslinker ratio of 20% w/v: 25% w/w hydrogel samples with this cross-linker strength at 90 × 90 µm as analyzed. The height parameters were obtained as root mean square height of 17.6 nm; mean peak height at 66.6 nm and mea pit height of 139 nm. For hydrogel with concentration of 25% w/v: 25% w/w, using cantilever tip scan window of 90 × 90 µm, heights parameters recorded were root mean
Fig. 5. Comparative plot showing the rheological behaviour of the hydrogels for two different compositions, S1 denotes for AAc: AAm 10% w/v: 25% w/w whereas S3 denotes 20% w/v: 25% w/w. The measurement was performed in strain sweep mode.
in the scale 10−3. The plot indicated as S1 in Fig. 5 showed that both elastic modulus (G′) and viscous modulus (G″) are independent of shearing time and it represents the stiffness of the hydrogel which is an indication of co-existence of both viscous and elastic properties. While for S3, it was observed that G′ increased over G″ by order one magnitude at low shearing time. But over sustained shearing, both elastic and viscous modulus assumed time independence. These characteristics properties could be beneficial for the consideration of hydrogel as a
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Fig. 6. Characteristics of surface topology measured through atomic force microscopy (AFM) on hy. (a) 15%w/w: 15%w/v; (b) 15%w/w: 20%w/v; (c) 15%w/w: 20%w/v; (d) 15%w/w: 25%w/v AAm: AAc Crosslinker Ratio.
bearing liquid film of the liquid drop causing the spreading of the liquid layer over the hydrogel surface (Hoffman, 2012). This property supports in-vivo cell culture in vascular networks that are hydrophilic in nature (Liu et al., 2013). The results presented in Table 2 showed the decreasing contact angles values, RMS of AFM for varying cross-linker ratio of the gelbased microchannel elastomer. The resulted showed that hydrogel matrix surface roughness decreases with increasing cross-linker ratio implying that hydrophilicity of the matrix increases with increasing concentration of acrylic acid in the polyacrylamide-based hydrogel (Enas, 2015; Farooqi et al., 2017). We observed that wetting property impacted by variation in acrylamide composition influences surface roughness and this could be implicated to the actions of intermolecular forces in the hydrogel matrix (Chakraborty et al., 2012).
Table 2 Comparison of wettability, surface roughness. Hydrogel specification (AAm_AAc ratio)
Contact angle (in degrees)
RMS height (from AFM measurements) µm
10% 15% 20% 25%
66 68 72 74
0.196 0.97 0.17 0.025
w/v_25% w/v_25% w/v_25% w/v_25%
w/w w/w w/w w/w
square height of 25.5 nm, mean peak height 150 nm and mean pit height of 51 nm. This value implies improved surface roughness on the hydrogel with increasing concentration of acrylic acid constituent of the polymeric hydrogel samples analyzed which is in consistent to literature report (see Hosseini et al., 2014b; Farahmand et al., 2015; Boks et al., 2008).
3.4. Comparative flow profiles of velocity distribution in hydrogel and PDMS micro-conduits
3.3. Effect of acrylamide composition of surface wettability A preliminary flow investigation was conducted to observe the behaviour of flow using hydrogel elastomer for selected flow rate. The behaviour of flow field at 1 µL/min for PDMS crosslinker ratio of 30:1 as captured in Fig. 7A showed the flow experienced very strong recirculation between channel axial lengths of 0.13–0.27 cm. An abrupt obstruction encountered by the flow was short-lived while strong recirculation energises the flow downstream the channel. Within the domain of flow, velocity magnitude which was high at the channel entry region suffered early decay probably due to the presence of unpolymerised gel in the flow domain (see Fig. 7B). However, recirculation occurred at about 0.1 unit length of the flow channel and velocity
Although hydrophilicity of the hydrogel surface could be assessed using the RMS technique, a more assuring test by determination of the surface contact angles were carried out as earlier discussed. Table 2 contained the contact angle values for varying acrylamide composition in the hydrogel. It could be observed that contact angle decreases with increasing acrylic acid content in the hydrogel matrix which is the hydrate constituent in the pre-polymer solution. It is also concluded that since contact angles were below 90 degree, that the surface is hydrophilic irrespective of the composition (Cha et al., 2013). There is thus a strong adhesive force between the hydrogel surface and the 222
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Fig. 7. A: Plot of velocity magnitude in PDMS microchannel of CR30:1 at flow rate of 1 µL/min. Fig. 7B: Velocity magnitude at flow rate of Q1 in 10% w/v: 25% w/w hydrogel microchannel at flow rate of 1 µL/min. Fig. 8. Comparative plot of flow velocity distribution in PDMS-hydrogel microchannel at 1 µL/min.
growth was rapid as the flow exit the PDMS-based microchannel. The comparative plots for both PDMS and Hydrogel at considerably near cross linker ratio presented in Fig. 8 showed that fluid flow in both microchannels assumed a similar profile. The maximum velocity attained was an order of magnitude lower in hydrogel microchannel. This could be alluded to the compliant nature of the hydrogel elastomer and could suggest a probable use to investigate flow where importance is attached to the flexible nature wall.
microchannels within the selected range of elastomer composition. Wettability of hydrogel elastomer shifts gradually within the hydrophilicity range with a mean deviation of 2.7 degree. Hydrogel elastomer is here presented as candidate elastomer for whole microchannel fabrication for studies where flexibility specifically fluid-channel flow compliance and hydrophilicity are of critical requirements.
4. Conclusion
This work was supported by the Department of Science and Technology of The Government of India and Federation of Indian Chamber of Commerce and Industries (FICI) through CV Raman International Fellowship awarded to the first author (Ebenezer Olubunmi Ige). The authors are grateful to Indian Institute of Technology Kharagpur for providing the facilities used in this research. Mr Sudipto Panda, Dr Shantimoy Kar and Prof. Sanatanu Chatophadhahy are well appreciated for their contribution to this work.
Acknowledgement
Hydrogel elastomer has been subjected to surface analysis to determine the visco-elastic, surface roughness and wettability behaviour under Newtonian fluid flow condition. The results obtained showed that storage modulus increases with increasing acrylamide composition which suggests improved structural rigidity when such elastomer is used as flow confinement in fluidic system and as replacement or mimic-agents of native biological conduits. Likewise, the visco-elastic response presents hydrogel as candidate elastomer for biomimetic studies of native flexible and compliant channels. The growth in surface roughness for morphology analysis with reducing acrylamide composition indicated the possibility of resistance to fluidic motion in
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