Development Of Separation Methods Research Articles

Preparation of imidazole-modified paper membrane for selective extraction of gallic acid and its structural and functional analogues from Pomegranate Peel.

In the search for pharmaceutically active compounds from natural products, it is crucial and challenging to develop separation methods that target not only structurally similar compounds but also a class of compounds with desired pharmaceutical functions. To achieve both structure-oriented and function-oriented selectivity, the choice of functional monomers with broad interactions or even biomimetic roles towards targeted compounds is essential. In this work, an imidazole (IM)-functionalized paper membrane was synthesized to realize selectivity. The IM was selected based on its capability to provide multiple interactions, participation in several bioprocesses, and experimental verification of adsorption performance. Using gallic acid as a representative component of Pomegranate Peel, the preparation conditions and extraction parameters were systematically investigated. The optimal membrane solid-phase extraction (MSPE) method allowed for enrichment of gallic acid from the complex matrix of Pomegranate Peel, enabling facile quantitative analysis with a limit of detection (LOD) of 0.1 ng mL-1. Furthermore, with the aid of cheminformatics, the extracted compounds were found to be similar in both their structures and pharmaceutical functions. This work offers a novel approach to preparing a readily synthesized extraction membrane capable of isolating compounds with similar structures and pharmaceutical effects, and provides an MSPE-based analytical method for natural products.

A Particular Separation Method Development and Validation of Nadolol and Bendroflumethiazide by using RP-HPLC

We have developed a simple, general, and reliable HPLC Method and validation for the simultaneous estimation of nadolol and bendroflumethiazide drugs according to (ICH) guidelines. Nadolol and bendroflumethiazide peaks have been observed at a retention time of 1.757, and 3.208 minutes, respectively, and they have kept 5 minutes as a total run time. As per linearity results, the average correlation coefficient of nadolol and bendroflumethiazide is 0.999 which indicates they have good linearity, robustness, and stability.

Chromatographic separation of silver-111 from neutron-irradiated palladium target: toward direct labeling of radiotracers

BackgroundSilver-111 is a promising β−-emitting radioisotope with ideal characteristics for targeted radionuclide therapy and associated single photon emission tomography imaging. Its decay properties closely resemble the clinically established lutetium-177, making it an attractive candidate for therapeutic applications. In addition, the clinical value of silver-111 is further enhanced by the existence of the positron-emitting counterpart silver-103, thus imparting a truly theranostic potential to this element. A so-fitting matching pair could potentially overcome the current limitations associated with the forced use of chemically different isotopes as imaging surrogates of lutetium-177, leading to more accurate and efficient diagnosis and treatment. However, the use of silver-111-based radiopharmaceuticals in vivo has faced obstacles due to the challenges related to its production and radiochemical separation from the target material. To address these issues, this study aims to implement a chromatographic separation methodology for the purification of reactor-produced silver-111. The ultimate goal is to achieve a ready-to-use formulation for the direct radiolabeling of tumour-seeking biomolecules.ResultsA two-step sequence chromatographic process was validated for cold Ag-Pd separation and then translated to the radioactive counterpart. Silver-111 was produced via the 110Pd(n,γ)111Pd nuclear reaction on a natural palladium target and the subsequent β−-decay of palladium-111. Silver-111 was chemically separated from the metallic target via the implemented chromatographic process by using commercially available LN and TK200 resins. The effectiveness of the separations was assessed by inductively coupled plasma optical emission spectroscopy and γ-spectrometry, respectively, and the Ag+ retrieval was afforded in pure water. Recovery of silver-111 was > 90% with a radionuclidic purity > 99% and a separation factor of around 4.21·10−4.ConclusionsThe developed separation method was suitable to obtain silver-111 with high molar activity in a ready-to-use water-based formulation that can be directly employed for the labeling of radiotracers. By successfully establishing a robust and efficient production and purification method for silver-111, this research paves the way for its wider application in targeted radionuclide therapy and precision imaging.

Cross-section measurements for 68Zn(p,2p)67Cu and 68Zn(p,2n)67Ga reactions using a newly developed separation method for the superposed gamma-ray spectra

We have developed a new analytical peak separation analysis for superposed gamma-ray peaks on ^{67}Cu and ^{67}Ga to measure the ^{68}Zn(p,2p)^{67}Cu and ^{68}Zn(p,2n)^{67}Ga reactions, unlike in most previous works that were employing a radiochemical separation to measure them. Based on the nuclear data such as the gamma-ray intensity and the half-life for each nuclide, we may develop a new analytical method that enables us to estimate the respective counts arising from each nuclide, thereby obtaining the nuclear reactions. The newly developed analytical method can universally be applied to separate the superposed gamma-ray spectra of any two nuclides, especially superior in separating the nuclides with different half-lives. In comparison with the data in the literature, the two reactions in the present work are in good agreement with those of some previous works. In addition, we compared the present ^{68}Zn(p,2n)^{67}Ga reaction without the peak separation to the data in the literature without the chemical separation, and find that a good agreement is evident, enhancing the reliability of the ^{68}Zn(p,x)^{65}Zn and ^{68}Zn(p,3n)^{66}Ga reactions, which are further measured in the present work

Stability-indicating normal-phase HPLC method development for separation and quantitative estimation of S-enantiomer of lacosamide in pharmaceutical drug substance and tablet dosage form.

Lacosamide (LA) is an antiepileptic medicine that is used to treat tonic-clonic seizures, partial-onset seizures, mental problems, and pain. A simple, effective, and reliable normal-phase liquid chromatographic technique was developed and validated to separate and estimate the enantiomer of (S-enantiomer) LA in pharmaceutical drug substance and drug product. Normal-phase LC was performed using USP L40 packing material (250 × 4.6mm, 5μm) and a mobile phase of n-hexane and ethanol at 1.0ml min-1 . The detection wavelength, column temperature, and injection volume used were 210 nm, 25°C, and 20 μl, respectively. The enantiomers (LA and S-enantiomer) were completely separated using a minimum resolution of 5.8 and accurately quantified without any interference in a 25-min run time. Accuracy study for stereoselective and enantiomeric purity trials was conducted between 10 and 200%, with recovery values ranging from 99.4 to 103.1% and linear regression values >0.997. The stability-indicating characteristics were assessed using forced degradation tests. The proposed normal-phase HPLC technique is an alternate approach to the official monograph methods (USP and Ph.Eur.) of LA, and it was successfully used in the evaluation of release and stability samples for both tablet dosage forms and pharmaceutical substances.

Boosting the Separation of Adeno-Associated Virus Capsid Proteins by Liquid Chromatography and Capillary Electrophoresis Approaches.

The purity of the three capsid proteins that make up recombinant adeno-associated virus (rAAV) is considered a critical quality attribute of gene therapy products. As such, there is a clear need to develop separation methods capable of rapidly characterizing these three viral proteins (VPs). In this study, the potential benefits and limitations of different electrophoretic and chromatographic methods were evaluated, including capillary electrophoresis-sodium dodecyl sulfate (CE-SDS), reversed phase liquid chromatography (RPLC), hydrophilic interaction chromatography (HILIC), and hydrophobic interaction chromatography (HIC), for the analysis of VPs obtained from different serotypes (i.e., AAV2, AAV5, AAV8, and AAV9). CE-SDS is considered to be the reference method and provides a suitable separation of VP1-3 proteins using generic conditions and laser induced fluorescence detection. However, the characterization of post-translational modifications (i.e., phosphorylation, oxidation) remains difficult, and species identification is almost impossible due to the lack of compatibility between CE-SDS and mass spectrometry (MS). In contrast, RPLC and HILIC were found to be less generic than CE-SDS and require tedious optimization of the gradient conditions for each AAV serotype. However, these two chromatographic approaches are inherently compatible with MS, and were shown to be particularly sensitive in detecting capsid protein variants resulting from different post-translational modifications. Finally, despite being non-denaturing, HIC offers disappointing performance for viral capsid proteins characterization.

Analytical Lifecycle Management (ALM) and Analytical Quality by Design (AQbD) based Analytical Method Development for Separation of Related Substances in Amodiaquine Hydrochloride along with its Degradation Products and Structural Elucidation by LC-Quadrupole-Time of Flight-Tandem Mass Spectrometry

The current study aimed to develop a robust, regulatory-flexible, stability-indicating ultra high performance liquid chromatography (UHPLC) analytical procedure compatible with mass spectrometry for the determination of impurities in amodiaquine hydrochloride using analytical lifecycle management (ALM) and analytical quality by design (AQbD) principles. The analytical target profile (ATP) and critical method attributes (CMAs) for the analytical method were identified. The pH of the mobile phase, flow rate and column oven temperature were identified as critical method parameters (CMPs) through risk assessment studies and they were screened and optimized using the design of experiments (DoE) to generate the design space (DS). Finally, all the impurities were well separated in a 15 min run time by using an Acquity UPLC BEH-C18 column with 20 mM ammonium acetate pH 8.0 as mobile phase A and acetonitrile as mobile phase B with a gradient program of time (min), % of B: 0/15, 1.0/15, 9/55, 12/55, 12.1/15 and 15/15. The flow rate was 0.3 mL min-1 and the column oven temperature were 30 ºC. Evaluated the robustness of the developed analytical method. Degradation products obtained from the forced degradation studies are well separated from process impurities and main compound. Based on the stress studies, impurity-3 was identified as a key degradation product in base degradation, while DP-1, impurity-3, DP-2, DP-3 and DP-4 were in the oxidative degradation. Total 9 impurities out of 4 process related and 5 degradation impurities were well separated in the developed method. Used the same method to LC coupled tandem mass spectrometer for separation and structure elucidation of degradation products. The method validation was carried out in accordance with ICH Q2 (R1) and USP <1225> guidelines.

Pyrolysis mechanism of natural fiber in cement-based composites at high temperatures

• A new method for fibre–matrix separation was developed; • The chemical composition change of the natural fibres in the cementitious matrix at different temperatures were investigated; • The main pyrolysis gas components of natural fibres at high temperatures were analyzed; • The pyrolysis kinetic parameters of natural fibres were determined based on the pyrolysis kinetic methods. The mechanism of pyrolysis of natural fibres in cementitious composites at high temperature has been investigated based on a new developed separation method. The influence of cementitious environment on the pyrolysis of natural fibres was revealed by examining the chemical composition, morphological changes and kinetic parameters of pyrolysis of natural fibres at different temperatures. The experimental results show that natural fibres are more susceptible to pyrolysis under high temperature conditions when embedded in a cementitious base. The fibre pyrolysis process can be considered to be through shrinkage, microfibre separation, fracture and eventually partial mass conversion into gas escape. Furthermore, the complex cementitious environment causes the crystallinity index ( CrI ) and crystallinity percentage ( % Cr 1 ) of natural fibres to decrease and then increase at elevated temperatures, which is different from the phenomena observed in a normothermic cementitious environment or under high temperature conditions alone. This work is of assistance and relevance to the evaluation of the durability of natural fibre reinforced cementitious composites at high temperature.

Method Development for Separation and Analysis of Tire and Road Wear Particles from Roadside Soil Samples

A comprehensive understanding of tire and road wear particles (TRWPs) and their detection and quantification in soils is still challenged by the lack of well-set standardized methods, inherent technological inconsistencies, and generalized protocols. Our protocol includes soil sampling, size separation, and organic matter removal by using hydrogen peroxide followed by density separation and analysis. In this context, roadside soil samples from different sites in Kansas and Ohio, USA, were collected and analyzed. Tire cryogrinds analogous to TRWPs were used to evaluate various density separation media, and collected particles more than 1 mm in size were then subjected to infrared spectroscopy (IR), thermogravimetric analysis (TGA), and scanning electron microscopy coupled with energy dispersive X-ray spectroscopy (SEM-EDX) to confirm TRWP presence. Particles smaller than 1 mm were Soxhlet extracted, followed by gas chromatography-mass spectrometry (GC-MS) to validate the presence of tire-related intermediates. SEM-EDX validated the presence of elemental combinations (S + Zn/Na) ± (Al, Ca, Mg, K, Si) attributed to tires. Ketones, carboxylic acids, epoxies, cyclohexane, and benzothiazole sulfenamide (BTS) intermediates were the most probable tire-related intermediates observed in the roadside soil samples. Thus, this simple, widely applicable, cost-effective sample preparation protocol for TRWP analysis can assist TRWP research advancement in terrestrial environments.

Development of methods for separation of imidacloprid and acetamiprid from biological liquids

Development of scientifically based forensic chemical and chemical-toxicological analysis of imidacloprid and acetamiprid poisoning cases. An extraction method was used to develop imidacloprid and acetamiprid in an acidified water method to separate them from blood and urine samples. In a 100 ml glass container, add 10 ml of test blood and 25 ml of urine and add 0.5 mg of working standard solutions of imidacloprid and acetamiprid. The model liquids were left to stir for 1 hour at room temperature, stirring occasionally. After the specified time, the pH of the model samples was checked dropwise from a 10% solution of sulfuric acid using a universal indicator and brought to 2.0–2.5 and left for 2 hours at room temperature. The mixture was then centrifuged at 3000 rpm, transferred to a centrifuge separator funnel and cleaned of ballast using 10 ml of organic solvent (imidacloprid–diethyl ether; acetamiprid–ethyl acetate). The pH of the aqueous part was brought to 9.0–9.5 using a 25% solution of ammonia, controlled using a universal indicator. The 3 were then extracted again with 10 ml of chloroform. 3–5 g of dehydrated sodium sulfate was filtered through a filter paper to remove moisture from chloroform extracts. The filtrates were dried at room temperature until a dry residue remained. The obtained dry residues were dissolved in 95% ethyl alcohol and qualitative and quantitative analyzes were performed. For the conduct of chemical-toxicological studies and forensic chemical examinations, an acidic aqueous method of separation of imidacloprid and acetamiprid from blood and urine samples was developed. For this, the liquid-liquid extraction method was used. High levels of imidacloprid and acetamiprid were obtained from biological fluids using the developed methods. A method has been developed to separate imidacloprid and acetamiprid from biofluids in blood and urine. The developed effective method of isolating pesticides was recommended for use in chemical-toxicological analyses.

Solvent-Impregnated Sorbents for Tantalum from Niobium Separation Using a Fixed-Bed Column.

Reactor-grade niobium steel is used as a construction material for nuclear reactors. In this case, the presence of tantalum, which is characterized by a 20 times higher active cross section for capturing thermal neutrons than the cross section of niobium (181Ta: 21.3 barn), cannot exceed 100 ppm. Analytical methods for quality and new separation method development control need very pure niobium matrices—niobium compounds with a low tantalum content, which are crucial for preparing matrix reference solutions or certified reference materials (CRMs). Therefore, in this paper, a new, efficient method for separating trace amounts of Ta(V) from Nb(V) using extraction chromatography with the use of sorbents impregnated with methyl isobutyl ketone MIBK solvent is proposed. Various types of MIBK-impregnated sorbents were used (AG® 1-X8 Anion Exchange Resin, AMBERLITE™ IRC120 Na Ion Exchange Resin, SERVACEL® Cellulose Anion Exchangers DEAE 52, active carbons of various grain size, carbonized blackcurrant pomace, carbonized chokeberry pomace, bentonite, and polyurethane foam in lumps). The highest tantalum removal efficiency was determined using active coal-based materials (>97%). The separation effectivity of tantalum from niobium was also determined in dynamic studies using a fixed-bed column with MIBK-impregnated active carbon. Solutions of various Nb:Ta weight ratios (1:1, 100:1, 1000:1) were used. The most impressive result was obtaining 70 mL of high purity niobium solution of tantalum content 0.027 ppm (in relation to Nb) with 88.4% yield of niobium from a solution of Nb:Ta, weight ratio 1000:1 (purge factor equaled 35,000). It proves the presented system to be applicable for preparation of pure niobium compounds with very low contents of tantalum.

Separation Sciences in Poland

This paper describes the most significant achievements related to the creation and development of separation methods. Tswiett Nerst’s achievements and the role of such centers as Lublin, Warsaw, Gdańsk, Poznań, Łódź, Silesia, Kraków and Toruń are presented, as well as the role of leaders who created theories and methodological studies on chromatography and related techniques. The role of sample preparation methods is emphasized based on the same or similar theoretical foundations. The directions of developing these techniques in Poland today and in the future are presented.

APPLICATION OF TARTARIC ACID DERIVATIVES IN ENANTIOSEPARATION OF RS-IBUPROFEN

A novel Tartaric acid derivative-based chiral resolving agent has been discovered for the enantiosepration of racemic Ibuprofen [(RS)-2-(4-(2-methyl propyl) phenyl) propanoic acid]. The processinvolves optimization of diastereomeric salt formed between (+) Ibuprofen and optically pure tartaric acid derivatives, followed by separating the individual enantiomer by acid salt preparation. Characterization of the synthesized compound has been carried out by elemental analysis, melting point, FT-IR, UV, HPLC, and 1H NMR spectra. The developed separation method is simple, cost-effective, and industrially viable.

Development of Supercritical Fluid Chromatographic Methods for Separation of Conazole Pesticide Enantiomers

Development of Supercritical Fluid Chromatographic Methods for Separation of Conazole Pesticide Enantiomers

Enantioresolution and Binding Affinity Studies on Human Serum Albumin: Recent Applications and Trends

The interaction between proteins and drugs or other bioactive compounds has been widely explored over the past years. Several methods for analysis of this phenomenon have been developed and improved. Nowadays, increasing attention is paid to innovative methods, such as high performance affinity liquid chromatography (HPALC) and affinity capillary electrophoresis (ACE), taking into account various advantages. Moreover, the development of separation methods for the analysis and resolution of chiral drugs has been an area of ongoing interest in analytical and medicinal chemistry research. In addition to bioaffinity binding studies, both HPALC and ACE al-low one to perform other type of analyses, namely, displacement studies and enantioseparation of racemic or enantiomeric mixtures. Actually, proteins used as chiral selectors in chromatographic and electrophoretic methods have unique enantioselective properties demonstrating suitability for the enantioseparation of a large variety of chiral drugs or other bioactive compounds. This review is mainly focused in chromatographic and electrophoretic methods using human serum albumin (HSA), the most abundant plasma protein, as chiral selector for binding affinity analysis and enantioresolution of drugs. For both analytical purposes, updated examples are presented to highlight recent applications and current trends.

Production of Fucoxanthin from Phaeodactylum tricornutum Using High Performance Countercurrent Chromatography Retaining Its FOXO3 Nuclear Translocation-Inducing Effect.

Phaeodactylum tricornutum is a rich source of fucoxanthin, a carotenoid with several health benefits. In the present study, high performance countercurrent chromatography (HPCCC) was used to isolate fucoxanthin from an extract of P. tricornutum. A multiple sequential injection HPCCC method was developed combining two elution modes (reverse phase and extrusion). The lower phase of a biphasic solvent system (n-heptane, ethyl acetate, ethanol and water, ratio 5/5/6/3, v/v/v/v) was used as the mobile phase, while the upper phase was the stationary phase. Ten consecutive sample injections (240 mg of extract each) were performed leading to the separation of 38 mg fucoxanthin with purity of 97% and a recovery of 98%. The process throughput was 0.189 g/h, while the efficiency per gram of fucoxanthin was 0.003 g/h. Environmental risk and general process evaluation factors were used for assessment of the developed separation method and compared with existing fucoxanthin liquid-liquid isolation methods. The isolated fucoxanthin retained its well-described ability to induce nuclear translocation of transcription factor FOXO3. Overall, the developed isolation method may represent a useful model to produce biologically active fucoxanthin from diatom biomass.

Application of the analytical quality by design principles to the development of a qualitative surface‐enhanced Raman scattering method: A proof of concept

AbstractAnalytical quality by design (AQbD) is a systematic approach that allows developing analytical methods in a more science‐based way, due to the higher quality of information collected during the development process. However, its application remains limited to the development of separation methods. (i.e., liquid chromatography and capillary electrophoresis). Its application to spectroscopic techniques remains underexplored, despite the potential benefits. The aim of this work was to demonstrate the application of the AQbD principles to surface‐enhanced Raman scattering (SERS) method development, detailing each step characterizing this approach. First, crystal violet (CV) was chosen as a model compound and the analytical target profile was defined. Risk assessment was performed to individuate the critical method parameters. The intensity of the CV band at 1175 cm−1 was selected as critical method attribute to be modeled and maximized. A screening study was conducted to study 11 aggregation agents (AAs) at different concentration levels (range 0.010–1 M) and in two solvent media (aqueous and methanolic). The two AAs exhibiting the strongest signal, one for each medium, were retained. Once these parameters were fixed, an I‐optimal design was applied to optimize the entire analysis process. Here, the volumes of nanoparticle suspension and AA solution added to the SERS samples were simultaneously varied. Two method operable design regions, one for each medium, were computed using a Bayesian design space approach. The latter enabled evaluating the failure risk associated to each experimental condition explored and choosing those that best fit the analytical purpose in the experimental domain.

Development of separation method for Alpha-Tocopherol and Gamma-Oryzanol extracted from rice bran oil using Three-Zone simulated moving bed process

• The chromatographic separation method of gamma-oryzanol and alpha-tocopherol was developed. • An environmentally friendly solvent was employed for the separation. • A three-zone simulated moving bed system was designed via process simulation. • The purities of greater than 98% for both compounds were suggested at the optimal condition. • The maximum productivity was obtained at the optimal condition. Gamma-oryzanol and alpha-tocopherol are bio-active compounds that can be found in crude extract of rice bran oil. With superior properties of gamma-oryzanol such as antioxidant activities, anticancer, and antidiabetic, while alpha-tocopherol is one type of vitamin-E, they are both widely used in foods, pharmaceuticals, nutraceuticals, and cosmetic. Therefore, the isolation of these compounds from the crude extract to obtain a single component with high purity is important. In this study, the separation of gamma-oryzanol and alpha-tocopherol using chromatographic method was investigated. The type of mobile phase for C18 column was selected based on the selectivity, back pressure, and toxicity. The adsorption of mono-component was performed to determine the adsorption parameters and the mathematical model for breakthrough curves was developed. The model and the associated parameters were validated by the adsorption experiments using binary mixture. The verified model was used in the design and optimization of three-zone simulated moving bed process. The mobile phase was the mixture of ethanol and water (90 :10 V/V%). At the optimal conditions of the three-zone simulated moving bed, the purity of extract product (rich in gamma-oryzanol), the purity of raffinate product (rich in alpha-tocopherol), and the separation productivity were 99.24%, 98.69% and 29.87 g/L⋅min, respectively.

Long-term changes in evapotranspiration over China and attribution to climatic drivers during 1980–2010

Evapotranspiration (ET) is one of the most important variables in terrestrial ecosystems, linking the carbon-water-energy cycles. In this study, we first analyze the spatial patterns of annual ET changes during 1980–2010 across China using four ET products: (i) the Global Land Evaporation Amsterdam Model version 3.0a (GLEAMv3.0), (ii) the EartH2Observe ensemble (EartH2Observe-En), (iii) the Global Land Data Assimilation System version 2.0 with Noah model (GLDAS2.0-Noah), and (iv) the Modern Era Retrospective-Analysis for Research and Application-Land (MERRA-Land). The results show that the spatial distribution of annual mean ET values and long-term changes derived from these four ET products are similar. Overall, large-scale increases in ET are observed in southeastern China, while decreases in ET over the northeast. Furthermore, we apply a newly developed separation method with the Budyko framework to quantify the individual contribution of five climatic factors to ET changes, including precipitation (P), net radiation (Rn), air temperature (T), vapour pressure deficit (VPD), and wind speed (u). It is found that the dynamics of P, Rn, and VPD are all strongly correlated with ET, suggesting that they are the major climatic factors influencing ET changes. Specifically, precipitation is the dominant factor for ET in water-limited regions, while ET changes in energy-limited regions are dominated by VPD according to all ET products except the EartH2Observe-En in which Rn and VPD have comparable performance. Our study highlights the importance of VPD in ET changes across energy-limited regions of China and suggests that the role of VPD in land surface-atmosphere interactions should be considered in future studies.

Pore Size Control via Multiple-Site Alkylation to Homogenize Sub-Nanoporous Covalent Organic Frameworks for Efficient Sieving of Xenon/Krypton.

Among various fission products generated in nuclear reactors, xenon and krypton are two important fission gases with high flow, diffusivity, and radioactivity. Moreover, xenon isolated from these products is an expensive industrial resource with wide applications in medicine and lighting, which makes the development of efficient methods for separation of xenon/krypton significant. However, it is usually difficult for xenon/krypton to be adsorbed by chemical adsorbents due to their inert gas properties, and sub-nanoporous adsorbents proven to be workable for the separation of xenon/krypton are still hard to prepare and regulate the pore size. Herein, we report two novel sub-nanoporous covalent organic frameworks (COFs), which were applied to the sieving of xenon/krypton for the first time. The sub-nanoporous COFs were synthesized via aldehyde-amine polycondensation reactions and the subsequent pore size regulation and homogenization process by using a facile, operational, and efficient multiple-site alkylation strategy. Impressively, the as-prepared sub-nanoporous COFs realized the efficient adsorption and sieving of xenon/krypton owing to their slightly larger pore sizes (∼7 Å) than the dynamic diameters of xenon/krypton and their larger pore volumes. The maximum adsorption capacity for xenon is up to 85.6 cm3/g, and the xenon/krypton selectivity can reach to 9.7. Moreover, the as-prepared COFs possess good γ-ray irradiation stability, which endows them with great potentials for the sieving of radioactive xenon/krypton in the practical application. The multiple-site alkylation strategy proposed in this study provides a valuable approach for the pore construction and control of the porous materials, especially the sub-nanoporous adsorption materials.