Spermine CE Phosphoramidite

Cat. No.: S1492359

Storage

Room temperature

Shipped In

Normal

Specifications

Condizioni di conservazione di stoccaggio

Room temperature

Spedito in

Normal

Application Protocols

The following is a general workflow for using a CE phosphoramidite modifier on an automated DNA/RNA synthesizer. Item‑specific settings are Not specified for this item; refer to CoA/Spec Sheet and your instrument’s manual.

Preparation:

• Equilibrate all reagents to room temperature in a dry environment.
• Dissolve the phosphoramidite at 0.05–0.1 M in anhydrous MeCN. Filter through a dry 0.2 µm PTFE filter into a moisture‑free reagent bottle. Blanket with argon and cap tightly.
• Load a compatible activator (e.g., 0.25–0.5 M ETT/BTT/DCI in MeCN). Ensure fresh capping and oxidation solutions.

Synthesis:

• Program the instrument for a modifier coupling step with extended coupling time (e.g., 5–15 min) or double coupling as needed.
• Proceed with standard detritylation, capping, and oxidation between steps per platform SOP.

Post‑synthesis:

• Cleave and deprotect using conditions compatible with the modifier’s protecting groups (e.g., NH4OH or AMA; temperature/time per platform).
• Purify by cartridge or HPLC. Adjust ion‑pairing strength to account for the polycationic spermine after deprotection.
• Analyze by LC–MS and analytical HPLC. Confirm mass shift corresponding to single modifier incorporation.

Storage of solutions:

• Use within several days if kept rigorously dry and inert; discard if significant hydrolysis is detected (drop in coupling performance, new peaks in 31P NMR/LC).

All steps are for research use only.

Biological Roles

This tab discusses the biological context of the spermine moiety incorporated by this phosphoramidite; it does not imply biological activity of the reagent itself.

• Spermine (literature): a ubiquitous endogenous polyamine present in prokaryotes and eukaryotes. It modulates nucleic acid structure, stabilizes compacted DNA, influences ribosomal function and translation fidelity, and participates in cellular stress responses. Intracellular spermine levels are tightly regulated by biosynthesis (ornithine decarboxylase pathway), transport, and catabolism (polyamine oxidases).
• Nucleic acid interactions: Spermine binds to the phosphate backbone through electrostatic interactions, affecting duplex stability, condensation, and higher‑order structures (e.g., triplexes, i‑motifs).
• Oligonucleotide modification rationale (literature): Introducing a spermine unit to synthetic oligos can modulate hybridization thermodynamics (often increasing apparent Tm for DNA/DNA or DNA/RNA duplexes in salt‑limited conditions), enhance binding to negatively charged biomolecules, and alter cellular trafficking of conjugates.
• Enzymatic tolerance: Polymerases and nucleases may be affected by proximal polycationic modifications; placement (5′ vs internal) can influence compatibility in downstream assays. Validation is recommended for each application.

Note: All uses of this reagent are for research only. Effects of spermine‑modified oligonucleotides are context‑dependent and should be empirically determined with appropriate controls.

Buffer Applications

This product is not a buffering agent. It is a phosphoramidite reagent for solid‑phase synthesis of modified oligonucleotides.

Practical guidance:

• After synthesis and deprotection, choose analysis/storage buffers appropriate for the modified oligo (e.g., water, 10 mM Tris‑HCl, pH 7.5–8.0, with/without 1–50 mM NaCl or ammonium acetate for MS).
• Polyamine‑modified oligos may interact more strongly with anionic buffer components; verify retention times and peak shapes when developing HPLC or CE methods.

For buffer recipes and pH ranges, refer instead to general oligonucleotide handling protocols; they depend on the final modified oligo, not on this reagent.

Green Alternatives

While CE phosphoramidite chemistry conventionally relies on acetonitrile and chlorinated solvents, greener strategies can reduce environmental impact without compromising performance.

Greener considerations (literature/general):

• Solvent optimization:
  • Replace part of DCM with 2‑MeTHF or CPME in washing steps where instrument compatibility allows; note that core coupling still typically requires dry MeCN for reliable kinetics.
  • Employ high‑purity recycled MeCN if validated for water/impurity content.
• Activators: ETT and BTT can offer faster couplings and higher per‑cycle efficiencies than tetrazole, potentially reducing reagent consumption and waste.
• Process intensification: Increase amidite concentration and reduce cycle time where validated to cut solvent per base added.
• Solid support selection: High‑loading, low‑swelling supports can decrease solvent use per cycle.
• Oxidation alternatives: Use aqueous tert‑butyl hydroperoxide or MeCN/H2O/I2 systems optimized to minimize halogenated waste; confirm compatibility with the modifier’s protecting groups.

Comparison (general; performance varies by platform):

• Conventional: MeCN + DCM workflow — robust, broad instrument compatibility, higher hazardous waste volume.
• Improved: MeCN‑centric with limited CPME/2‑MeTHF substitution — reduced chlorinated solvent use, similar performance, requires validation of seals and drying.

Caveats:

• Moisture control is paramount; many “green” solvents have higher water affinity and require rigorous drying.
• Any substitution should be qualified on the intended synthesizer with small‑scale trials and analytical confirmation (HPLC/MS).

Pharmaceutical Uses

No excipient or pharmacopeial status is provided for this item. It is supplied strictly for research and laboratory use.

Context for professionals (general/literature):

• Role in development: Spermine CE phosphoramidite can be used in discovery‑stage synthesis of modified oligonucleotides to study structure–activity relationships, delivery, and target engagement.
• Manufacturing R&D: In process development for therapeutic oligonucleotides, polyamine modifiers are screened for effects on purification, stability, and conjugation workflows. Any use beyond research requires rigorous vendor qualification and regulatory assessment; this listing does not provide GMP documentation.
• Analytical characterization: Modified oligos are typically qualified by LC–MS, ion‑pair or HILIC HPLC, UV/Vis, and capillary gel electrophoresis. Additional assays (residual solvents, counter‑ion content) are application‑specific.

Item‑specific notes:

• Grade/Purity: Not specified for this item; refer to CoA/Spec Sheet.
• Regulatory status: Not specified for this item; refer to CoA/Spec Sheet.

No therapeutic or clinical claims are made. Any translational use requires independent validation and compliance with applicable regulations.

Physical Properties

Item‑specific values are not provided in the Product Data and should be confirmed from the CoA/Spec Sheet.

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Purity/grade: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular weight: Not specified for this item; refer to CoA/Spec Sheet.
• Melting/boiling point: Not specified for this item; refer to CoA/Spec Sheet.
• Density, refractive index, UV cutoff: Not specified for this item; refer to CoA/Spec Sheet.

General/literature characteristics of CE phosphoramidites (for context only):

• Physical state: typically low‑melting solids, gums, or viscous oils depending on linker/protecting groups (literature).
• Solubility: highly soluble in anhydrous acetonitrile (MeCN), dichloromethane (DCM), THF, and to some extent in DMF; poorly soluble in protic solvents and water (literature).
• Stability: moisture‑sensitive; hydrolyzes to the corresponding H‑phosphonate/ phosphate derivatives; air oxidation to phosphate can occur; best handled under dry inert atmosphere (literature).
• PKa/logP: not routinely defined due to protected multifunctional structure; the protected polyamine is non‑basic until deprotection (literature).

Practical notes for use (general):

• Prepare stock solutions freshly (commonly 0.05–0.1 M in dry MeCN) and protect from moisture.
• Pass solutions through a dry 0.2 µm PTFE filter before loading to synthesizers to minimize particulates.
• Use desiccated delivery lines and bottles to preserve activity.

Quality and Grades

• Grade/Purity (item‑specific): Not specified for this item; refer to CoA/Spec Sheet.

How to interpret quality for CE phosphoramidites (general guidance for professional users):

• Coupling performance: High‑quality phosphoramidites exhibit high coupling efficiencies on DNA/RNA synthesizers (often >98% per step under optimized conditions; literature). Performance is best assessed by trityl cation release monitoring, crude oligo HPLC, and mass spectrometry.
• Assay/identity tests commonly expected on CoA: 31P NMR (single major P(III) resonance), 1H/13C NMR, LC–MS, HPLC purity, water content (Karl Fischer), residual activator/solvent profile. Item‑specific numeric limits are not provided here and should be taken from the CoA/Spec Sheet.
• Stabilizers: Some amidites are supplied with trace stabilizers (e.g., BHT) or as solutions in MeCN with added base scavengers. For this item, stabilizers are Not specified for this item; refer to CoA/Spec Sheet. Stabilizers can influence background during synthesis and may require blank runs.
• Particle/flow characteristics: For instrument compatibility, low particulate levels and controlled viscosity are important; filtration through 0.2 µm PTFE is recommended prior to loading.
• Storage verification: On receipt, verify intact seal, dryness (no crystallization or phase separation for solutions), and clear labeling. Retain CoA/SDS lot records for GMP‑adjacent research environments.

Note: When substituting amidite lots/suppliers, re‑optimize coupling time and activator concentration to maintain crude purity and yield.

Reaction and Applications

Primary application: installation of a spermine polyamine moiety onto synthetic oligonucleotides via solid‑phase phosphoramidite chemistry.

Key reactions and workflow (literature/practice):

• Coupling: The 2‑cyanoethyl N,N‑diisopropyl phosphoramidite couples to the 5′‑OH (or suitable internal hydroxyl) of a solid‑supported oligo in the presence of an activator (e.g., ETT, BTT, DCI). Spermine CE amidites may require extended coupling times relative to standard nucleoside amidites.
• Oxidation: Post‑coupling, P(III) is oxidized to phosphate using I2/H2O/pyridine/THF mixtures; sulfurization can be used if a phosphorothioate linkage is desired at the attachment site (if compatible with protecting groups).
• Capping: Unreacted 5′‑OH are capped (Ac2O/NMI) to suppress n−1 sequences.
• Detritylation: If a DMT‑type protecting group is present on the modifier handle, it is removed with acid (e.g., 3% TCA in DCM) to continue chain elongation.
• Deprotection/cleavage: After synthesis, standard base treatment (e.g., NH4OH or AMA) removes base‑labile protections and releases the oligo; spermine protecting groups are typically base‑labile and are removed during this step (specific conditions should follow the supplier’s recommendations).

Why add spermine? (biophysical/literature):

• Spermine imparts a localized polycationic character that can increase affinity for nucleic acids, modulate Tm, and influence cellular uptake of conjugates. It is explored in antisense, siRNA, and probe design to modulate hybridization and delivery properties.

Use cases:

• 5′ terminal modification of DNA/RNA, internal modification via aminolinkers, conjugation sites for polyamine‑mediated binding studies.

Reaction Conditions

The following are general literature/practice conditions for CE phosphoramidite couplings; optimize for your synthesizer and sequence. Item‑specific parameters are not provided here.

• Stock solution: 0.05–0.1 M in anhydrous MeCN; for challenging incorporations, up to 0.15–0.2 M if solubility allows. Prepare fresh and keep dry/inert.
• Activators (typical):
  • ETT (0.25–0.5 M in MeCN)
  • BTT (0.25–0.5 M in MeCN)
  • DCI (0.25–0.5 M in MeCN) Coupling times for bulky modifiers: 5–15 min; some platforms benefit from double couplings.
• Oxidation: I2 (0.02–0.05 M) in THF/pyridine/H2O/MeCN mixtures for 30–90 s per cycle. Alternative sulfurization if required (e.g., PADS, DDTT) depending on desired linkage and protecting group compatibility.
• Capping: Cap A (Ac2O in THF) / Cap B (NMI/DMAP in THF/pyridine) for 30–60 s.
• Detritylation: 3% TCA in DCM (or DCM/MeCN) for 30–90 s; monitor to avoid overexposure that can impact sensitive modifiers.
• Cleavage/deprotection (general): Concentrated NH4OH (55–65 °C, 2–8 h) or AMA (1:1 MeNH2 in EtOH / NH4OH, 65 °C, 5–30 min) are common. Ensure compatibility with spermine protecting groups for complete deprotection; consult supplier documentation.
• Workup/purification: Desalting by cartridge, SEC, or HPLC (ion‑pair reversed‑phase or HILIC). Polyamine‑modified oligos may elute earlier/later depending on ionic strength; method scouting is recommended.

Quality checks: UV trityl assay (if applicable), analytical HPLC, ESI‑MS or MALDI‑MS, and, when relevant, CE.

Safety and Handling

Authoritative safety details for this item are Not specified for this item; refer to CoA/Spec Sheet and SDS.

• GHS classification, pictograms, signal word, H‑statements: Not specified for this item; refer to SDS.

General safety information for CE phosphoramidites (literature/practice):

• Hazards: Typically cause skin/eye irritation; harmful if inhaled or ingested. Moisture‑sensitive; reaction with water liberates amines/alcohols and may form irritating vapors. Oxidation state changes possible upon air exposure.
• Incompatibilities: Avoid moisture, acids, strong oxidizers, and protic solvents during handling. Activators (e.g., tetrazoles) increase reactivity—avoid contact with oxidizers/strong bases outside controlled synthesis.
• PPE: Laboratory coat, nitrile gloves, splash goggles. Handle in a fume hood when weighing, dissolving, or transferring.
• Handling: Use dry syringes/vials, anhydrous solvents, and inert gas blanket (Ar/N2). Cap containers promptly.
• First aid (overview; follow SDS):
  • Skin: Remove contaminated clothing; wash with soap/water. Seek medical advice if irritation persists.
  • Eyes: Rinse cautiously with water for several minutes; remove contact lenses; seek medical attention.
  • Inhalation: Move to fresh air; seek medical attention if symptoms occur.
  • Ingestion: Rinse mouth; do not induce vomiting; seek medical attention.
• Spill response: Absorb with inert material (silica, vermiculite), keep dry, collect for disposal as organic waste.

Always defer to the item’s SDS and institutional EHS policies for definitive guidance.

Solvent Selection

This product is a moisture‑sensitive CE phosphoramidite intended for automated oligonucleotide synthesis. Solvent choice governs reactivity and stability.

General/literature solvent guidance:

• Primary solvent: Anhydrous acetonitrile (MeCN) is the industry standard for dissolving and delivering phosphoramidites (high polarity, low viscosity, low nucleophilicity). Typical use: 0.05–0.1 M solutions.
• Alternative solvents: Dichloromethane (DCM) or THF can be used for stock preparation of certain hydrophobic modifiers; DMF can aid solubilization of more polar amidites. Always ensure anhydrous grade (<50 ppm H2O) and instrument/material compatibility.
• Activator solutions: ETT, BTT, DCI, or 5‑ethylthio‑1H‑tetrazole in MeCN enhance coupling kinetics (literature).
• Incompatibilities: Avoid alcohols, water, and other protic solvents; they rapidly quench the P(III) species. Avoid amine‑rich solvents that can prematurely deprotect protecting groups.

Selection tips (practice):

• If solution shows haze or slow delivery, switch from MeCN to a MeCN/DCM (9:1 to 7:3) blend to improve solubility (validate instrument seals for chlorinated solvents).
• For high‑load modifier couplings (internal/5′), use higher amidite concentration (up to 0.15–0.2 M) in MeCN if viscosity permits; otherwise employ a small fraction of DCM or THF.
• Filter freshly prepared solutions through dry 0.2 µm PTFE and sparge with dry argon to minimize dissolved moisture.

Storage and Reconstitution

Item‑specific conditions:

• Storage Conditions: Room temperature (per Product Data).
• Shipped in: Normal (per Product Data).

Best practices for CE phosphoramidites (general/literature):

• Keep tightly sealed in a dry, inert atmosphere (argon/nitrogen) with desiccant. Moisture rapidly degrades P(III) amidites.
• Protect from prolonged exposure to air and light. If supplied as a solid/gum, minimize open‑air handling; if supplied as a solution, verify dryness before connecting to the synthesizer.
• Do not refrigerate/freezer‑store unless specified on the CoA, as repeated warming can cause condensation; instead, maintain a consistently dry ambient environment.

Reconstitution (for use):

• Dissolve in anhydrous acetonitrile to the desired concentration (commonly 0.05–0.1 M; higher if needed and soluble).
• Optional: Add a small fraction of DCM or THF to aid solubility for viscous modifiers, ensuring instrument compatibility.
• Filter through a dry 0.2 µm PTFE membrane into a moisture‑free bottle; sparge with dry argon and cap.

In‑use stability:

• Monitor performance via trityl cation release (if applicable) and crude HPLC. Declining coupling efficiency often indicates hydrolysis; prepare fresh solution.

Unspecified parameters (appearance, shelf life, exact stabilizers, water limits): Not specified for this item; refer to CoA/Spec Sheet.

Research Use: For research use only (per Product Data).

Structure and Identity

Spermine CE Phosphoramidite is a solid‑phase DNA/RNA synthesis building block that installs a spermine (polyamine) moiety via a standard 2‑cyanoethyl N,N‑diisopropyl phosphoramidite group.

• Product name: Spermine CE Phosphoramidite (modifier for oligonucleotide synthesis)
• SKU: S1492359
• CAS: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular weight: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES: Not specified for this item; refer to CoA/Spec Sheet.
• InChIKey: Not specified for this item; refer to CoA/Spec Sheet.
• Structural features (general/literature):
  • Contains a terminal 2‑cyanoethyl N,N‑diisopropyl phosphoramidite functional group for automated coupling (literature).
  • The cargo is a spermine polyamine (–NH–(CH2)3–NH–(CH2)4–NH–(CH2)3–NH2) typically protected on amines with base‑labile groups during synthesis (literature).
  • A linker or spacer connects spermine to the phosphoramidite handle; many commercial designs also include a DMT (4,4′‑dimethoxytrityl) or other protecting group on the attachment site (literature).
• 2D description (general): a central protected polyamine chain attached through a linker to a phosphoramidite bearing a diisopropylamino substituent and a 2‑cyanoethyl group; the P center is trivalent and chiral but used as a racemic reagent (literature).

Notes: Exact structure, protecting group pattern, and linker length can vary by supplier/model. For definitive identity parameters (formula, MW, spectra), consult the item’s CoA/Spec Sheet.

Synthetic Utility

Functional role: A P(III) 2‑cyanoethyl N,N‑diisopropyl phosphoramidite used to append a protected spermine polyamine to oligonucleotides on solid support.

Key synthetic features (literature/practice):

• Chemoselective activation: Formation of a highly reactive phosphoramidite–tetrazolide intermediate enables rapid coupling to the terminal 5′‑OH (or internal hydroxyls on modified supports).
• Orthogonal protection: The polyamine nitrogens are typically masked with base‑labile groups, surviving acidic detritylation and iodine oxidation, then unveiled during global deprotection.
• Modularity: Compatible with DNA and RNA synthesis cycles; can be introduced at 5′, internal (using appropriate aminolink/diol handles), or at the 3′ end when immobilized on specialized supports.
• Post‑synthetic diversification: The introduced spermine can serve as a polyvalent handle for electrostatic assembly, crosslinking (with diacids/activated esters after deprotection), or further conjugation through selectively addressable sites if present in the linker design.
• Retrosynthetic value: Treat as a “late‑stage modifier” allowing fine tuning of oligo properties without altering the base sequence.

Analytical control:

• Monitor coupling by on‑instrument trityl cation release (if DMT‑protected) and verify incorporation by LC–MS of the crude and purified oligo.
• Be mindful of multiple charge states during MS due to polycationic spermine after deprotection; ammonium acetate buffers facilitate desalted ESI‑MS analysis.

Target Specificity

Not applicable. This product is a chemical building block (phosphoramidite) and does not have biological target specificity, antigen binding, or clone/isotype attributes.

For sequence‑specific or target‑binding behavior, evaluate the final spermine‑modified oligonucleotide in its intended assay context.