Electronic Supporting Material
forMicrochimicaActa
Immobilization of zirconium-glycerolatenanowireson magnetic nanoparticlesforextraction of urinary ribonucleosides
Jing Xua, Zheng Zhanga, b, Xiao-Mei Hea, Ren-Qi Wanga, c, Dilshad Hussaina, d, Yu-Qi Fenga*
a Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Department of Chemistry, Wuhan University, Wuhan 430072, China
b State Key Laboratory of Proteomics, Beijing Proteome Research Center, BeijingInstitute of Radiation Medicine, Beijing 102206, China
c College of Chemistry and Chemical Engineering, Lanzhou University, Lanzhou 730000, China
d Division of Analytical Chemistry, Institute of Chemical Sciences, BahauddinZakariya University Multan (60800), Pakistan.
S.1 Instrumentation and analytical conditions
Morphologies of Fe3O4@ZrGly and ZrGly were characterized with a transmission electron microscope (TEM, JEOL, Kyoto, Japan). Element compositions were determined by EDX-720 energy-dispersive X-ray analysis (EDX, Shimadzu, Kyoto, Japan).Attenuated total reflectance- Fourier transform infrared spectra of dried materials were determined by Thermo Nicolet AVTAR-360 (ATR-FTIR, Thermo Fisher Scientific, Madison, USA). Thermal gravity analyses were performed with a TG-DTA 6300 thermal analyzer (TGA, PerkinElmer, MA, USA). Magnetization curves were obtained from with a PPMS-9 vibrating sample magnetometer (VSM, Quantum Design, San Diego, USA). Nitrogen adsorption measurements were performed at 77K using a Micromeritics ASAP 2020 surface area and pore size analyzer (Micromeritics, GA, USA). The composites were activated in vacuum and heated to 393K for 4 h to remove any physically adsorbed substances before analysis.The BET (Brunauer-Emmett-Teller) specific surface areas were obtained from the adsorption branch in relative pressure range 0.05–0.30. BJH (Barrett–Joyner–Halenda) model was used to estimate the pore distribution from the desorption branch. The 13C NMR spectra were recorded on a Bruker Advanced II (400 MHz, WI, USA) spectrometer with d6-DMSO as the solvent and tetramethylsilane (TMS) as an internal reference. The solid-state CP/MAS13C NMR spectra were obtained on a Bruker Avance IIIHD 400WB NMRspectrometer (100.56 MHz, WI, USA), with a 90degree 1H-pulsewidth of4.0 usand a contact time of 2 ms, spinning at the magic angle at 10 kHz frequency. The13Cchemicalshiftsweredeterminedusingsolidexternalreference,hexamethylbenzene(HMB).Themethyl groupsofHMBresonatedat17.35 ppmrelative totetramethylsilane(TMS).
The HPLC-UV experiments were carried out on LC-20A (Shimadzu, Japan) HPLC system. A Hisep C18-T column (250 mm ×4.6 mm i.d., 5 μm, Weltech Co., Ltd., Wuhan, China) was used for HPLC separation. 2mMammonium hydrogen carbonate (solvent A) and methanol (solvent B) were employed as mobile phases at a flow rate of 1 mL/min. A gradient of 0-8 min 5% B, 8-20 min 5-20% B, 20-27.5 min 20-80% B, 27.5-30 min 80% B and 30-40 min 5% B was used. For all the HPLC–UV experiments, the detection wavelength was set at 254 nm and the injection volume was 10 μL.
S.2 TGA of Fe3O4 and Fe3O4@ZrGly
The composition of Fe3O4@ZrGly was confirmed by TGA analysis (Fig. S3). The first weight loss at 50-150 oC was attributed to the removal of adsorbed water and the second weight loss between 200-350 oCwasduetothe decomposition of glycerol moiety bonded to zirconium[1]. Theseresults indicated that the content of glycerol moiety in Fe3O4@ZrGly was about 35%. The weight loss of Fe3O4@ZrGly between 200-350 oC suggested a large amount of ZrGlywas immobilized on Fe3O4. Besides, an extra weight loss in the temperature range of 500-600 oC was observed which wasdue to the dehydration of zirconium oxyhydrate.
S.3Adsorptin of 2'-deoxycytidine
For the evaluation of the hydrophilic property of Fe3O4@ZrGly toward deoxyribonucleosides, 2'-deoxycytidine was chosen as a hydrophilic probe. 5 mg of Fe3O4@ZrGly was added to 500 μLof loading buffer (2µg/mLfor 2'-deoxycytidine, dissolved in 1% FA), and the mixture was vortex mixed at room temperature for 5 min.Then the supernatant was collected for HPLC-UV analysis.
S.4Binding constant calculation
Scatchard plots were also used to calculate binding constant of the probe with the material. According to Scatchard method, the data should fitto the following equation (1):
(1)
Q is the analyte absorbed on the sorbent,Qmax is the theoretical maximum adsorption of analyte on the sorbent, [C*] is the concentration of theanalyte in solution, and Kdis the desorption constant. According to the above rule, a plot of Q/[C*]versus Q should yield a straight line if only one type of adsorption site exists[2]. Such a case was observed when adsorbing with Fe3O4@ZrGly (1:106) and the desorption constants Kdof Fe3O4@ZrGly (1:106) was calculated as 41 mg/mL.
S.5Extraction optimization
Fe3O4@ZrGly could isolateribonucleosides from aqueous samples because of theircoordination interactionswithZrGly nanowires. Major factors, such as loadingsolution and effect of salt concentrations, were optimizedusing nucleosides standard samples (2 μg/mL) as analytes.Since nucleosides are either weak acidic nucleosides or weak alkaline nucleosides, pH has a greatinfluence on molecular status of nucleosides, which can affect theextraction efficiency of the sorbents dramatically. As shown in Fig. S7a, 0.5% NH3·H2Ois optimal for binding four nucleosides. To demonstrate the salt interferenceeffect in 0.5% NH3·H2O, the standard samples of nucleosides were extracted by adding different concentrations of NaCl. No change in extraction efficiencyof Fe3O4@ZrGly was observed in the presence of salt (Fig. S7b), which suggested that Fe3O4@ZrGly had good salt tolerancetill salt concentration was 80 mM. As innormal urine the concentrations of salt are not more than 75 mmol·day-1[3,4], which is lower than that of our added, materials salts tolerance is endurable for the atrocious circumstances.
Fig.S1TEM graphs of (a) Fe3O4, (b) ZrGly and (c) Fe3O4@ZrGly, and SEM graphs of (d) Fe3O4, (e) ZrGly and (d)Fe3O4@ZrGly.
Fig.S2 ATR-FTIR patterns of Fe3O4, Fe3O4@ZrGly, ZrGly and glycerol*.
*The two negative peaks were the absorption peak of CO2
Fig.S3TGA spectra of Fe3O4 and Fe3O4@ZrGly.
Fig.S4 LC-UV chromatograms of 2'-deoxycytidine standard solutions before (a) and after (b) enrichmentwith Fe3O4@ZrGly. The amount of 2'-deoxycytidine was 1 μg.
Fig.S513C-NMR spectra of substance resolved of ZrGly in 10 vt % TFA aqueous solution.
Fig. S6The adsorption equilibrium plot, the adsorption isotherm curves based on Freundlich mode and Langmuir mode in the adsorption process of Fe3O4@ZrGly, commercial ZrO2 and CeO2 towards adenosine.
Fig. S7The maximum adsorption capacities of Fe3O4@ZrGlys with different molar ratios of zirconium acetylacetonate and glycerol.
Fig. S8 Optimization of the MSPE parameters. The effect of loading buffer (a) and salt concentrations (b) on the extraction efficiency.
Table S1.Effects of synthesis parameters on the atom content of Fe3O4@ZrGly with characterization by EDS.
Materials / C (At%) / O (At%) / Fe (At%) / Zr (At%)ZrGly (1:106) / 67.3 / 29.8 / - / 3.0
Fe3O4@ZrGly (1:53) / 15.9 / 52.8 / 30.5 / 0.8
Fe3O4@ZrGly (1:106) / 29.8 / 47.8 / 20.2 / 2.2
Fe3O4@ZrGly (1:264) / 28.7 / 42.4 / 27.2 / 1.7
Fe3O4@ZrGly (1:432) / 20.7 / 50.9 / 26.7 / 1.7
*The numbers within brackets represent the molar ratio of zirconium acetylacetonate and glycerol.
Analytes / Linear range (μg·mL-1) / Calibration curve data / LOD(ng·mL-1) / LOQ
(ng·mL-1)
Slope
(×104) / Intercept / R2
Adenosine / 0.05-10.0 / 5.55 / 0.05 / 0.9938 / 1.7 / 5.1
Cytidine / 0.05-10.0 / 2.35 / 0.01 / 0.9971 / 9.2 / 28
Guanosine / 0.05-10.0 / 1.60 / -0.03 / 0.9938 / 19 / 58
Uridine / 0.05-10.0 / 0.60 / 0.03 / 0.9974 / 8.0 / 24
Table S2.Linearity characteristics of four nucleosides in diluted urine.
Table S3. Precision and recoveries of four nucleosides from spiked human urine.
Analytes / Intra-day(RSD, %, n=5) / Inter-day
(RSD, %, n=3) / Recovery
(%, n=3)
0.2a / 5.0 / 10.0 / 0.2 / 5.0 / 10.0 / 0.2 / 5.0 / 10.0
Adenosine / 7.4 / 4.6 / 2.7 / 11.2 / 6.0 / 3.8 / 107
±11.9 / 115
±6.9 / 112
±4.2
Cytidine / 9.5 / 3.4 / 2.4 / 7.8 / 5.2 / 2.9 / 100
±7.8 / 100
±5.2 / 98.8
±2.9
Guanosine / 9.5 / 9.4 / 5.9 / 5.5 / 4.2 / 4.3 / 106
±5.9 / 103
±4.3 / 90.6
±3.9
Uridine / 12.4 / 2.6 / 3.6 / 2.4 / 1.8 / 2.3 / 109
±2.6 / 112
±2.0 / 113
±2.6
aSpiked concentration (μg·mL-1)
Table S4. An overview on recently reported nanomaterial-based methods for preconcentration of nucleosides
Analytical technique / Adsorbent / Adsorbentamount (mg) / Adsorption capacity
(mg·g-1) / LOD
(ng·mL-1) / Ref.
DSPE-
HPLC-MS / Boronic acid-functionalized magnetic attapulgite / 10 / 14 / 2-41 / [5]
SPE-
HPLC-UV / Boronic acid-functionalized magnesia-zirconia / 500 / 124 / 3-10 / [6]
DSPE-
HPLC-UV / ATTA@MPS@PBA@C12mimBr / 20 / 31 / 0.06-0.46 / [7]
DSPE-
LC-MS/MS / Titania−zirconia nanoparticles coated on porous silica spheres / 5 / 35 / 2.9-52 / [8]
SPME-
nESI-MS/MS / Zirconia nanocoating cellulose paper / 10 / - / 0.01–1.26 / [9]
SPE-
HPLC-UV / Binary boronic acid-functionalized attapulgite / 50 / 20 / 4-17 / [10]
In-tip SPE-
HPLC-UV. / Polyethyleneimine-modified boronate affinity fibrous cotton / 10 / 0.7 / 3.5-4.7 / [11]
SPME- HPLC-UV / Zirconium-doped magnetic microspheres / 50 / 38 / 5-17 / [12]
SPME- HPLC-UV / Boronic acid derivativefunctionalized magnetic polysulfone capsules containing mesoporous silicananoparticles / 40 / - / 1.1-1.7 / [13]
SPME- HPLC-UV / Filter paper modified withphenylboronic acidfunctionalizedmesoporous silica / 17.6 / 10 / 2-11 / [14]
SPME- HPLC-UV / Fe3O4@ZrGly / 20 / 36 / 1.7-19 / This work
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