Stabilization Reagent Research Articles

GC×GC-TOFMS Analysis of Fecal Metabolome Stabilized Using an At-Home Stool Collection Device

Stool is a mixture of excrement, microbiota, enzymes, undigested material, and small molecules. Fecal metabolomics has gained interest recently, owing to advances in metabolomics and growing research into both the host’s physiology and the gut microbiome. One challenge with fecal metabolomics is preserving the sample integrity from collection until analysis, as the microbiota and enzymes continue to alter the metabolome following defecation. Currently, flash-freezing or lyophilization are utilized to minimize post-collection metabolome changes; however, this requires complex equipment and immediate processing, precluding the possibility for at-home sampling. Commercial devices containing stabilizing solvents have been developed to facilitate at-home collection, ambient transport, and sample storage. Here, we explore the efficacy of a commercially available stool collection device with a stabilization reagent tailored to fecal metabolomics. Stool samples from six donors were either processed shortly post-collection or stored at room temperature for seven days in the tube, with and without the stabilization reagent. Comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-TOFMS)-based untargeted metabolomics was utilized for analyzing extracted metabolites. Chemometric analysis was used to evaluate the performance of the device. We found that the device with the stabilization reagent minimized changes in the metabolite profile relative to unstabilized stool left at room temperature for one week.

Characteristics of RNA Stabilizer RNApro for Peripheral Blood Collection.

Peripheral blood is the most practical tissue for human immune system gene expression profiling because it is easily accessible, whereas the site of primary infection in certain diseases may not be easily accessible. Due to the ex vivo instability of RNA transcripts, a key challenge in the gene expression analysis of blood samples is the rapid sample handling and stabilization of the mRNA by adding an RNA preservative (PAXgeneTM Blood RNA Tubes, TempusTM Blood RNA tubes, RNAlater Stabilization Reagent, RNAgard® Blood Tubes). BioMole (Turin, Italy) has developed a novel blood stabilizer, called RNApro, in which RNA is stabilized during phlebotomy and sample storage. In this study, RNApro performance intended as RNA yield, integrity, and stability was evaluated. Our results show that blood samples stored at -80 °C and re-extracted after 7 years show no differences in terms of quantity, quality, and amplificability. The samples in the RNAlater stabilization solution can be stored at room temperature for up to one week or at 4 °C for up to one month. Similar results can also be observed for PAXgene tubes, Tempus tubes, and RNAgard tubes. In agreement with these data, the RNApro stabilization solution preserves the RNA from degradation for up to 1 month at 4 °C and 1 week at room temperature. RNApro can be stored indifferently at -80, -20, 4 °C, or room temperature for up to 2 months after, and then could be stored at -80 °C for up to seven years. In summary, our study is the first to analyze the performance of an RNA stabilizer called RNApro. We can conclude that several studies have shown significant differences in gene expression analysis when the sample was preserved in different RNA stabilizers. Therefore, it is desirable to standardize the method of nucleic acid conservation when comparing data from transcriptomic analyses.

An Innovative Method to Generate Bromine Monochloride for Trace Hg Analysis.

This work presents a new method for generating a BrCl solution, starting from the commercially available dibromodimethylhydantoin (DBDMH). This method is notable due to the straightforward, safe and clean performance, being based on a simple addition of DBDMH into aqueous HCl. The whole procedure is finished in about 20min. An advantage of the proposed method is avoiding of tedious reagents pre-cleaning by prolonged thermal treatment, spontaneous overheating and excessive vapor evolution. The resulting BrCl stabilization reagent is low enough in mercury content to be directly used in trace mercury analysis. A thorough study of the BrCl solutions prepared by this method showed that they conform in all respects to US EPA 1631e/2002 requirements.

Methodological considerations for aqueous environmental RNA collection, preservation, and extraction.

Environmental RNA (eRNA) analysis is expected to infer species' physiological information (health status, developmental stage, and environmental stress response) and their distribution and composition more correctly than environmental DNA (eDNA) analysis. With the prospect of such eRNA applications, there is an increasing need for technological development for efficient eRNA detection because of its physicochemical instability. The present study conducted a series of aquarium experiments using zebrafish (Danio rerio) and validated the methodologies for capture, preservation, and extraction of eRNA in a water sample. In the eRNA extraction experiment, an approximately 1.5-fold increase in lysis buffer volume resulted in a more than sixfold increase in target eRNA concentration. In the eRNA capture experiment, although GF/F and GF/A filters yielded similar eRNA concentrations, a GF/A filter may be capable of passing through more volume of water samples and consequently collecting more eRNA particles, given the time required for water filtration. In the eRNA preservation experiment, the use of RNA stabilization reagent (RNAlater) allowed for stably preserving target eRNA on a filter sample at -20 and even 4°C for 6days at least. Altogether, the findings enable the improvement of eRNA availability from the field and easily preserve eRNA samples without deep-freezing, which will contribute to the refinement of eRNA analysis for biological and physiological monitoring in aquatic ecosystems.

Abstract 5605: Ambient storage and stabilization of proteins and cell-free nucleic acids in whole blood samples

Abstract Introduction: Highly anticipated advancements in biotechnology and instrumentation now allows for simultaneous detection of multiple biomarkers. This is accelerating the cancer research field towards the development of multianalyte-based in vitro diagnostic tests for early detection and/or monitoring of cancer. These multianalyte liquid biopsy tests are often based on detection of cancer-related proteins and/or tumor-specific genomic and transcriptomic changes (circulating cell-free nucleic acids) in plasma. A specialized and proprietary stabilization reagent has been developed for use in a blood collection tube with the capability to simultaneously stabilize protein biomarkers of interest and cell-free nucleic acids. Methods: Blood samples from self-proclaimed healthy donors were drawn into evacuated blood collection tubes containing a new stabilization reagent and stored at room temperature. Plasma was isolated using a general double-spin protocol (1800 × g for 15 min and 2800 × g for 15 min) and frozen at -80 °C until use. Plasma levels of proteins of interest including cancer markers and immuno-oncology checkpoint markers were measured by Ella Simple Plex Assays (Bio-Techne) and Luminex xMAP® Technology Assays (Bio-Techne, samples tested by LuminexPLORE Lab). Total cell-free nucleic acids were isolated using the QIAamp Circulating Nucleic Acid Kit and the resultant cfDNA or cfRNA was analyzed with fluorometric-based assay (quantitative) and Bioanalyzer/Tapestation (qualitative). Results: Compared with blood samples collected in EDTA and ACD-A tubes, the new stabilization reagent maintained the draw time levels of plasma proteins of interest such as TGF-β1, MMP-9, IL-8/CXCL8, CCL5/RANTES, Granzyme B, CEA and PD-L1 for up to 5 days of room temperature storage in whole blood. The reagent also effectively maintained draw time levels of plasma cell-free nucleic acids to near draw time levels. In addition, the new reagent also minimized in vitro hemolysis during room temperature storage. Conclusion: This new and breakthrough stabilization reagent simultaneously maintains the draw time levels of proteins of interest and cell-free nucleic acids for an extended period of time in whole blood. It provides cancer researchers and clinical assay developers confidence in blood sample integrity and flexibility in sample storage, transport, and processing at ambient temperature for multianalyte analysis or multiomics profiling. Citation Format: Jing Li, Lisa Bartron, Jordan LaRue, Sama Mehta, Nicholas M. George. Ambient storage and stabilization of proteins and cell-free nucleic acids in whole blood samples. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5605.

Remediation of the lead–zinc composite-contaminated soil using stabilization reagent in an industrial region of Nanjing, China

Remediation of the lead–zinc composite-contaminated soil using stabilization reagent in an industrial region of Nanjing, China

Evaluation of a bacterial group 1 LEA protein as an enzyme protectant from stress-induced inactivation.

Late embryogenesis abundant (LEA) proteins are hydrophilic proteins that lack a well-ordered tertiary structure and accumulate to high levels in response to water deficit, in organisms such as plants, fungi, and bacteria. The mechanisms proposed to protect cellular structures and enzymes are water replacement, ion sequestering, and membrane stabilization. The activity of some proteins has a limited shelf-life due to instability that can be caused by their structure or the presence of a stress condition that limits their activity; several LEA proteins have been shown to behave as cryoprotectants in vitro. Here, we report a group1 LEA from Azotobacter vinelandii AvLEA1, capable of conferring protection to lactate dehydrogenase, catechol dioxygenase, and Baylase peroxidase against freeze-thaw treatments, desiccation, and oxidative damage, making AvLEA a promising biological stabilizer reagent. This is the first evidence of protection provided by this LEA on enzymes with biotechnological potential, such as dioxygenase and peroxidase under in vitro stress conditions. Our results suggest that AvLEA could act as a molecular chaperone, or a "molecular shield," preventing either dissociation or antiaggregation, or as a radical scavenger, thus preventing damage to these target enzymes during induced stress. KEY POINTS: • This work expands the basic knowledge of the less-known bacterial LEA proteins and their in vitro protection potential. • AvLEA is a bacterial protein that confers in vitro protection to three enzymes with different characteristics and oligomeric arrangement. • The use of AvLEA as a stabilizer agent could be further explored using dioxygenase and peroxidase in bioremediation treatments. AvLEA1 protects against freeze-thaw treatments, desiccation, and oxidative damage on three different enzymes with biotechnological potential.

Extraction of Noncondensed Lignin from Poplar Sawdusts with p-Toluenesulfonic Acid and Ethanol.

The traditional pretreatment leads to the recalcitration of C-C bonds during lignin fractionation, thus hindering their depolymerization into aromatic monomers. It is essential to develop an applicable approach to extract noncondensed lignin for its high-value applications. In this work, noncondensed lignins were extracted from poplar sawdust using recyclable p-toluenesulfonic acid for cleaving lignin-carbohydrate complex bonds effectively and ethanol as a stabilization reagent to inhibit lignin condensation. Lignin yield of 83.74% was recovered by 3 mol/L acid in ethanol at 85 °C for 5 h, and carbohydrates were well preserved (retaining 98.97% cellulose and 50.01% hemicelluloses). During lignin fractionation, the acid concentration and extraction time were the major drivers of condensation. Ethanol reacted with lignin at the α-position to prevent the formation of the condensed structure. The extracted lignin depolymerized over the Pd/C catalysts gave a yield of 50.35% of aromatic monomers, suggesting that the novel extraction process provided a promising way for noncondensed lignin production.

Importance of experimental information (metadata) for archived sequence data: case of specific gene bias due to lag time between sample harvest and RNA protection in RNA sequencing.

Large volumes of high-throughput sequencing data have been submitted to the Sequencing Read Archive (SRA). The lack of experimental metadata associated with the data makes reuse and understanding data quality very difficult. In the case of RNA sequencing (RNA-Seq), which reveals the presence and quantity of RNA in a biological sample at any moment, it is necessary to consider that gene expression responds over a short time interval (several seconds to a few minutes) in many organisms. Therefore, to isolate RNA that accurately reflects the transcriptome at the point of harvest, raw biological samples should be processed by freezing in liquid nitrogen, immersing in RNA stabilization reagent or lysing and homogenizing in RNA lysis buffer containing guanidine thiocyanate as soon as possible. As the number of samples handled simultaneously increases, the time until the RNA is protected can increase. Here, to evaluate the effect of different lag times in RNA protection on RNA-Seq data, we harvested CHO-S cells after 3, 5, 6, and 7 days of cultivation, added RNA lysis buffer in a time course of 15, 30, 45, and 60 min after harvest, and conducted RNA-Seq. These RNA samples showed high RNA integrity number (RIN) values indicating non-degraded RNA, and sequence data from libraries prepared with these RNA samples was of high quality according to FastQC. We observed that, at the same cultivation day, global trends of gene expression were similar across the time course of addition of RNA lysis buffer; however, the expression of some genes was significantly different between the time-course samples of the same cultivation day; most of these differentially expressed genes were related to apoptosis. We conclude that the time lag between sample harvest and RNA protection influences gene expression of specific genes. It is, therefore, necessary to know not only RIN values of RNA and the quality of the sequence data but also how the experiment was performed when acquiring RNA-Seq data from the database.

Issue Highlights — May 2021

Issue Highlights — May 2021

Trailblazing Next Generation Sodium Battery Chemistry through Tunable Sodium Phosphorothioates

Next-generation battery chemistry based on earth-abundant elements of sodium and sulfur can deliver a high theoretical specific energy of 1274 Wh kg-1, making low cost battery technologies possible. Nevertheless, the formation of unanchored soluble intermediate sulfur species and the precipitation of low electronically/ionically conductive solids impede the realization of such systems. Herein, by introducing another naturally plentiful element of phosphorus, we invented a new series of sodium phosphorothioates by taking advantage of the chemical affinity among these three elements, which solves the abovementioned challenges. These new materials not only serve as a high capacity cathode but also function as a stabilization reagent for sodium anode, contributing to a novel battery chemistry that delivers high electrochemical potentials and capacity. As a proof of concept, we demonstrate a high-performance sodium battery, showing an initial capacity delivery of 440 mAh g-1 along with a cyclic retention of 80% for over 400 cycles. The superiority of this new battery chemistry is fundamentally investigated by both experimental characterizations and theoretical calculations. This study can lay the groundwork in designing a low cost, high performance sodium battery chemistry that potentially meets the grid-scale energy storage requirements beyond the current lithium ion technologies. Figure 1

Rapid and Cost-Effective RNA Extraction of Rat Pancreatic Tissue.

Regardless of the extraction method, optimized RNA extraction of tissues and cell lines are carried out in four stages: 1) homogenization, 2) effective denaturation of proteins from RNA, 3) ribonuclease inactivation, and 4) removal of contamination from DNA, proteins, and carbohydrates. However, it is very laborious to maintain the integrity of RNA when there are high levels of RNase in the tissue. Spontaneous autolysis makes it very difficult to extract RNA from pancreatic tissue without damaging it. Thus, a practical RNA extraction method is needed to maintain the integrity of pancreatic tissues during the extraction process. An experimental and comparative study of existing protocols was carried out by obtaining 20-30 mg of rat pancreatic tissues in less than 2 minutes and extracting the RNA. The results were assessed by electrophoresis. The experiments were carried out three times for generalization of the results. Immersing pancreatic tissue in RNA stabilization reagent at -80 °C for 24 h yielded high integrity RNA, when the RNA extraction reagent was used as the reagent. The results obtained were comparable to the results obtained from commercial kits with spin column bindings.

Amphiphilic cellulose and surfactant mixtures as green frothers in mineral flotation. 1. Characterization of interfacial and foam stabilization properties

The stabilization of the froth phase is among the most important aspects for efficient separation in flotation. While the behavior of froth stabilization reagents has been thoroughly studied, there have only been minimal changes to their formulations in the past decades. Simultaneously, there is an increasing need for both more efficient processes due to declining ore grades and more sustainable methods due to increased environmental awareness.The current commercially applied frothers consist mainly of aliphatic alcohols and oligomeric glycol ethers. The present manuscript proposes a profound change to the state-of-art frother formulations by introducing the first ever detailed interfacial characterization of an aqueous polymer-surfactant (PS) mixture consisting of a nonionic amphiphilic polymer and a nonionic surfactant pair to be used as a flotation frother. The polymeric component of the mixture is a biodegradable cellulose derivative, namely hydroxypropyl methyl cellulose (HPMC), produced from sustainable sources. In the first part of this series, the dynamic surface tension, bubble size distribution and foam stability were measured for PS-mixtures with various compositions and concentrations. This work shows that the interaction of the components in this unique PS-system results in significant improvements to foam formation behavior compared with commercial frothers, including: i) a synergistic decrease of surface tension; ii) accelerated diffusion of surfactants to the air-liquid interface; iii) bubble size reduction; and iv) a self-stabilizing foam phase.

Distribution characteristics and comparison of chemical stabilization ways of heavy metals from MSW incineration fly ashes

With the purpose of fully understanding the potential toxicity of heavy metals of various particle sizes in fly ash (FA) and finding an optimal chemical reagent stabilization scheme for fly ash with single or mixture reagents, basic physicochemical characteristics of FA was investigated using multiple chemical stabilization reagent schemes to stabilize heavy metals. The following compounds were used to develop single and mixed chemical stabilization schemes for heavy metals in waste incineration FA: sodium sulfide (Na2S), sodium dihydrogen phosphate (NaH2PO4), ammonium dibutyl dithiophosphate (DDTP), and 2,4,6-tri-mercapto-S-triazine trisodium salt (TMT-15). The results showed that the various particle sizes of FA were mainly distributed in the range of 48–1700 μm. FA was packed with heavy metals, and the smaller the particle size, the more toxic it was. The speciation distribution of Cu, Zn, Pb, Cd, and Cr in different particle sizes was identical to that of the mixed samples. In the experiments of single reagent stabilization scheme, 8% Na2S, 8% NaH2PO4, 4.2% DDTP or 4.2% TMT-15 could decrease the leaching concentration to meet the limits set by Chinese law, but 8% Na2S adding scheme had the lowest cost among them. However, in the mixture reagent scheme, 1.2% Na2S, 1.2% NaH2PO4 and 0.8% DDTP was more effective and even cheaper than other mixture or single schemes.

Comparison of three methods to stabilize bronchoalveolar lavage cells for flowcytometric analysis.

Flowcytometric analysis of lymphocytes and their subpopulations in bronchoalveolar lavages (BAL) can support the diagnosis of interstitial lung diseases. This analysis should be done within 4hr after lavage due to rapid cell deterioration. We tested three methods in order to stabilize for at least 28 days the BAL cell populations to allow delayed flowcytometric analysis in order to facilitate external quality assurance (EQA). We compared an in-house, dual-step stabilization method for BAL cells with results of two different commercial available stabilization reagents: TransFix® and Streck Cell Preservative™. All three methods were compared with native BAL cells as reference. BAL samples from six patients were tested on six occasions following stabilization from 1 to 28 days by flow cytometry. Following stabilization and storage at 4°C, BAL cell suspensions had stable light scatter patterns and lymphocyte subsets. As expected, rapid deterioration of cells was seen with native BAL cells. The stabilized lavages showed more stable counts of WBC and lymphocyte populations with only minor differences found between the three methods. If analysis of the BAL cells is performed more than 24 hr after the lavage, stabilized BAL cells are superior to native cells. The in-house method can be used for EQA purposes with stability for at least 28 days. The TransFix and Streck methods might be useful for postponed diagnostic analysis of lavage cells but did not meet our 28 days criterion defined needed for EQA purposes.

Sensitive determination of hydrogen peroxide in real water samples by high spin peroxo complex.

In this paper, a fast, cheap, simple, sensitive and selective spectrophotometric method based on high spin peroxo-Fe(III)-EDTA complex in the alkaline medium was developed for the determination of hydrogen peroxide (H2O2) in real water samples. The purple-coloured complex with a maximum absorbance at a wavelength of 525 nm was formed. Various parameters such as type of stabilizer reagent and its concentration, reaction time, Fe(III), EDTA and NH3 concentration were optimized. The method was confirmed with the Beer’s law with a molar absorption coefficient of 267.36 L mol-1 cm-1 in the 8.3 ×10-6 –4.08 ×10-3 mol/L concentration range. Sandell’s sensitivity of the proposed method was also calculated as 0.188 μg/cm2 . LOD and LOQ were determined as 2.5 ×10-6 and 8.3 ×10-6 mol/L, respectively. Intraday and interday relative standard deviation of the proposed method for 2.0 ×10-4 mol / L of H2O2 were found as 1.5% and 6.1%, respectively. The developed method is suitable for fast monitoring of H2O2 in different types of aqueous water samples without any sample preparation steps and acceptable recovery values between 90% and 118% were obtained. In the sample analysis, H2O2 removed solutions from the real water samples were used for blank correction in their analysis and this process provides more reliable and accurate results in real sample analysis.

Unlocking the potential of sustainable chemicals in mineral processing: Improving sphalerite flotation using amphiphilic cellulose and frother mixtures

The transition towards cleaner production of primary minerals demands technologies that improve productivity while lowering their environmental impact. The present study explores a novel approach to support this transition by partially substituting a commercial froth stabilization reagent in mineral froth flotation (DowFroth200) with an amphiphilic cellulose derivative (namely, hydroxypropyl methyl cellulose) that is nontoxic, biodegradable and can be produced from sustainable sources. The performance of the cellulose-frother mixture is explored using sphalerite as case study.In the first place, the interaction between the two frother molecules was studied from the perspective of two-phase foam characteristics. The use of the polymer-surfactant (PS) mixture studied reported an increased foam stability and small bubble sizes, attributed to a combination of liquid film drainage prevention and steric effects of cellulose macromolecules. As a result, when used as frothers in the flotation of sphalerite, the PS-mixture exhibited a robust behavior resulting in advantages such as: i) improved separation efficiencies; ii) consistent recovery values, even under highly alkaline pH, iii) maintaining high recoveries when the collector dosage was reduced down to 25% of typical values used in the industry; and iv) faster flotation kinetics. The results obtained aim to demonstrate that properly designed green chemical systems can have a positive impact in the mining sector.

HCB dechlorination combined with heavy metals immobilization in MSWI fly ash by using n-Al/CaO dispersion mixture

Municipal solid waste incineration fly ash (MSWI-FA) has been strictly controlled as hazardous waste globally because it contains various heavy metals and dioxins. This study prepared a nanometallic Al/CaO (n-Al/CaO) dispersion mixture via ball-milling as a reductive stabilization reagent for the simultaneous immobilization of heavy metals and detoxification of POPs like substance in MSWI-FA. Under optimal conditions, Cu, Zn, Cd, Cr, Ni, and Pb had been significantly immobilized (over 99.9 %) and the leaching concentration of Zn, Cd, Cr, Ni, and Pb were below the detectable limit. Simultaneously, 82.43 % of HCB can be destructed into alkanes and amorphous carbon. The porous structure of the fly ash and alkaline surface of n-Al/CaO promoted the adsorption and cracking of HCB. The highly active n-Al/CaO interacted with water as the hydrogen donor to promote the reductive dechlorination process. Hydrocalumite was a new mineral formed from the adsorption and complexation of heavy metal. Therefore, n-Al/CaO can strengthen the control of heavy metals in the S/S treatment of MSWI-FA, effectively detoxify chlorinated organics, and reduce environmental health risks.

Improvement in RNA quantity and quality in cervico-vaginal cytology

The separation of exfoliated cells from the brushes used during cervico-vaginal smears is difficult, a problem which may affect the quality of ribonucleic acid (RNA) extracted. We compared the results of RNA extraction from cervico-vaginal cytology samples according to the type of tubes, preservative solutions, and storage temperature. The samples included exfoliated cervico-vaginal cytological specimens from patients with human papilloma virus 16, positive for cervical intraepithelial neoplasia or cervical cancer. Exfoliated cells were obtained by shaking a brush in a conventional rigid vial tube or squeezing the brush in a soft vial tube. RNA quantity and quality were compared between the two tubes. The concentration and purity of RNA (A260/A280 and A260/A230 ratios) was compared amongst five groups: Group 1, standard frozen storage; Group 2–4, RNA stabilization reagents with room temperature [RNAlater RNA Stabilization Reagent, RNAprotect cell Reagent and AllProtect Tissue Reagent]; and Group 5, Surepath Preservative fluid. To demonstrate the utility of the extracted RNA for PCR-based cDNA synthesis, GAPDH and E6 were targeted and gel band densities of GAPDH and E6 were measured. The median RNA concentration was significantly higher in the soft tubes compared with the rigid tubes (100.2 vs. 7.1 ng/μL, p = 0.0209). The purity of the RNA was higher in soft vial tubes than in rigid vials, as measured by A260/280 and A260/230 ratios. The RNA concentration, purity, and GAPDH density of groups 1, 2 and 3 were significantly higher than those of groups 4 and 5. Moreover, E6 density of group 1 and 2 was significantly higher than that of group 3, 4 and 5. The use of soft tubes enhanced the mRNA quantity and quality in cervico-vaginal cytology. The products of mRNA extraction using RNAlater RNA Stabilization Reagent and RNAprotect Cell Reagent at room temperature were comparable to those obtained by conventional frozen storage. Our protocol improved the yield and quality of RNA and might produce better results for molecular analysis in cervico-vaginal cytology.

Fast Pyrolysis of Organosolv Lignin: Effect of Adding Stabilization Reagents to the Extraction Process

Lignin is extracted by an improved organosolv method in the presence of different stabilization reagents including formaldehyde, formic acid, and methanol. The structure and thermochemical properties of the extracted lignin were characterized by gel permeation chromatography, Fourier transform infrared spectroscopy, heteronuclear single quantum correlation nuclear magnetic resonance, and thermogravimetric analysis techniques. During the extraction pretreatment, the fragmentation of lignin was suppressed via the addition of formaldehyde. The characteristic structure of natural lignin was retained by inhibiting the β-O-4 bond from cleavage. The fast pyrolysis of the extracted lignin was analyzed by the pyrolysis-gas chromatography/mass spectrometry technique. Compared with the formaldehyde-stabilized lignin, unstabilized lignin appeared to be more thermally resistant to volatilization. Because of the stabilization effect of formaldehyde, the pyrolysis yield of condensable volatiles was tripled; meanwhile, l...