Rifampicin-d8

Cat. No.: R757906

Storage

Room temperature

Specifications

Storage

Room temperature

Application Protocols

Example protocol (research LC–MS/MS quantitation; adapt to your method):

• Materials: Rifampicin‑d8 (internal standard), rifampicin analyte, DMSO (for stock), ACN/MeOH, water (LC–MS grade), formic acid or ammonium formate, calibration matrix (e.g., stripped plasma), SPE cartridges (optional).

• Stock and working solutions:

  1. Prepare rifampicin‑d8 stock at 1–10 mg/mL in DMSO (amber vial). Store aliquots.
  2. Prepare working IS at 50–500 ng/mL in ACN:water (50:50) with 0.1% formic acid.

• Calibration/QC standards:

  • Spike analyte into matrix across expected range; add a fixed concentration of rifampicin‑d8 to all calibrators, QCs, and unknowns prior to extraction.

• Sample preparation (protein precipitation):

  1. Aliquot 50–100 µL matrix into tubes.
  2. Add 3–5 volumes of cold ACN containing IS (vortex 30 s).
  3. Centrifuge (≥12,000×g, 10 min); transfer supernatant to amber vials.

• LC–MS/MS conditions (illustrative):

  • Column: C18, 2.1×50 mm, 1.7–3 µm.
  • Mobile A: water + 0.1% formic acid (or 5 mM ammonium formate); Mobile B: ACN or MeOH.
  • Gradient: start 30% B → 90% B over 3–5 min; flow 0.3–0.5 mL/min.
  • ESI+: monitor rifampicin transition(s) and +8 Da IS transitions that retain deuterium.

• Acceptance: Evaluate accuracy (85–115%), precision (CV ≤15%), matrix factor with IS‑normalization, carryover, and short‑term/long‑term stability per institutional SOPs.

Note: Parameters above are general guidance based on literature and common practice, not product‑specific specifications.

Biological Roles

Context (literature; research use only): Rifampicin is a member of the rifamycin (ansamycin) class that binds the bacterial DNA‑dependent RNA polymerase β‑subunit, sterically blocking elongation of nascent RNA. Deuteration (rifampicin‑d8) preserves the parent’s biochemical interactions while enabling mass‑spectrometric discrimination for tracer and quantitation work.

• Molecular interaction (literature):

  • Rifampicin occupies the RNA exit channel in RNAP, forming hydrogen bonds and hydrophobic contacts with conserved residues; activity depends on the phenolic/hydrazone functionalities and the ansa bridge conformation.
  • Resistance mutations cluster near the rifampicin‑binding pocket (rpoB gene), which is relevant for interpreting analytical studies on mutant strains in research settings.

• Physicochemical determinants:

  • Amphoteric molecule (phenols and piperazine) with pH‑dependent charge distribution influencing permeability and protein binding.
  • Strong plasma protein binding in biological systems (literature), which affects extraction recoveries during bioanalytical method development.

• Isotopic labeling rationale:

  • d8 internal standards co‑elute with the analyte and exhibit near‑identical extraction/ionization efficiencies, enabling accurate correction for matrix effects, ion suppression, and recovery losses.

• Application domains (research only):

  • Pharmacokinetic/pharmacometabolomic method development in preclinical models.
  • Stability and degradation pathway elucidation (photo‑/hydrolytic products) using isotope tracking.

No therapeutic or clinical use is implied for this material; it is intended strictly for laboratory research and analytical applications.

Buffer Applications

Rifampicin‑d8 is not a buffering agent and is not used to establish or maintain pH in biochemical assays. Instead, it is typically dissolved in organic solvents (e.g., DMSO, methanol, acetonitrile) and then diluted into existing buffered mobile phases or extraction solutions.

• Practical guidance (analytical context):
  • Common LC–MS mobile phases: water with 0.1% formic acid or volatile ammonium buffers (formate/acetate/bicarbonate, 2–20 mM) paired with acetonitrile or methanol. Choose pH to balance analyte stability and chromatographic retention.
  • Avoid strong basic buffers (>pH 9) or strong acids (<pH 2) for extended periods, as rifamycins can undergo hydrolysis/oxidation (literature).

For pH control in assays, select an appropriate buffer system (e.g., phosphate, HEPES, ammonium formate) independent of this compound.

Green Alternatives

Although Rifampicin‑d8 itself is not a solvent or reagent used in bulk, greener practices can be applied to its handling and analytical use.

• Solvent choice and reduction:

  • Prefer acetonitrile over halogenated solvents for extractions; both ACN and MeOH are commonly used and have established waste streams with lower environmental impact than chlorinated solvents.
  • Minimize DMSO volumes by preparing concentrated stocks and micro‑aliquoting to reduce solvent waste and container rinsing.

• Process intensification:

  • Use microextraction (SPE microcartridges, SPME fibers) to cut organic solvent consumption during sample prep.
  • Adopt UHPLC with short columns and higher flow efficiencies to reduce mobile‑phase use.

• Stabilization vs. waste:

  • Protect from light to prevent degradation, which otherwise generates additional waste during re‑preparation of standards and QC samples.

• Comparative guidance (general):

  • Traditional: Liquid–liquid extraction with large volumes of ethyl acetate/chloroform.
  • Greener alternative: Protein precipitation with ACN or MeOH, followed by SPE cleanup using water/ACN mobile phases; eliminates chlorinated solvents and reduces volume.

• Waste management:

  • Collect antibiotic‑containing waste separately when possible and treat as hazardous organic waste per institutional policy; avoid discharge to drains to limit environmental antibiotic load.

These measures maintain analytical performance while lowering environmental footprint without altering the compound itself.

Pharmaceutical Uses

This product is supplied for research use only. Rifampicin‑d8 is not an active pharmaceutical ingredient or excipient for administration. In a pharmaceutical R&D and quality environment, isotope‑labeled standards like rifampicin‑d8 are used as analytical tools.

• Typical roles in pharma labs (non‑clinical):

  • Internal standard for LC–MS/MS quantitation of rifampicin in preclinical formulations, dissolution samples, and process intermediates.
  • Support for method development/validation: selectivity, accuracy, precision, recovery, matrix effects, and stability studies as per bioanalytical guidance.
  • System suitability and ongoing performance checks in QC of dosage‑form extracts (research/developmental stages).

• Standards and compendia (context):

  • Pharmacopeial monographs commonly exist for rifampicin (parent), not for deuterated analogues; d‑labeled materials are nonetheless widely accepted for internal standardization when isotopic purity and identity are documented.

• Formulation considerations (analytical):

  • Stock solutions are typically prepared in DMSO, MeOH, or ACN and spiked into sample diluents compatible with the chromatographic method.

No claims of therapeutic efficacy, safety, or clinical diagnostic utility are made or implied for this item.

Physical Properties

Item-specific specifications (this product):

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Melting point, boiling point, density, refractive index, water/solvent content, UV cutoff, metal limits: Not specified for this item; refer to CoA/Spec Sheet.

Literature/computed properties (context; not product specifications):

• Parent rifampicin average MW: ~822.94 g/mol; rifampicin‑d8 theoretical average MW ~831.0 g/mol (computed by +8.06 u; exact value depends on labeling).
• Physical state: typically an orange to red‑orange crystalline solid/powder (literature for rifampicin). Color arises from the naphthohydroquinone chromophore.
• Solubility (literature):
  • Water: very low at neutral pH due to zwitterionic/aggregative behavior.
  • Organic: soluble in DMSO and DMF; moderately soluble in methanol, ethanol, acetonitrile (solubility increases with small amounts of base or acid).
• pKa (literature, parent): multiple ionizations; reported pKa values ca. 1–2 (phenolic), ~7–8 (piperazine), and higher for secondary alcohols; overall amphoteric behavior.
• logP/logD (literature, parent): logP typically reported ~3–4; logD strongly pH‑dependent due to ionizable piperazine and phenols.
• UV–vis (literature): strong absorbance in the visible (λmax around 475–500 nm, solvent/pH‑dependent) useful for HPLC/LC‑MS UV tracking.
• Stability notes (literature): sensitive to light (photoisomerization/oxidation) and to hydrolysis/oxidation at extreme pH; more stable in the dark, under inert atmosphere, and in aprotic solvents.

Practical note: For quantitative LC–MS, prepare concentrated DMSO stocks (e.g., 1–10 mg/mL), then dilute into aqueous/organic mobile phase with 0.1% formic acid or ammonium buffer immediately before use to minimize degradation.

Quality and Grades

• Item-specific grade/purity: Not specified for this item; refer to CoA/Spec Sheet.

• What to expect for isotope‑labeled analytical standards (general guidance):

  • Chemical purity: Typically ≥95% for small‑molecule internal standards; higher purities facilitate clean chromatograms and stable calibration curves. Verify by HPLC/LC–MS trace on CoA.
  • Isotopic enrichment: For “d8” materials, deuterium incorporation is often ≥98 atom% at specified positions; residual unlabeled or partially labeled isotopologues should be reported on the CoA because they affect isotopic overlap and correction factors.
  • Positional labeling: The exact deuteration sites in rifampicin‑d8 determine retention‑time matching and fragmentation pathways in MS/MS; consult the Spec Sheet for positional map.

• Stabilizers/inhibitors: Not specified for this item; refer to CoA/Spec Sheet. If present, stabilizers (e.g., ascorbate, BHT) can influence UV‑vis and MS baselines; account for them in blanks.

• Fit for purpose:

  • LC–MS/MS internal standard for quantitation of rifampicin in complex matrices (biofluids, environmental, formulation extracts). Deuterium labeling yields co‑elution with minimal isotope effect, aiding matrix‑matched quantitation.
  • For photolabile analytes, suppliers may recommend amber glass and inert‑gas backfill; absence/presence should be confirmed on receipt.

• Documentation: Request CoA/SDS for batch‑specific purity, isotopic distribution, residual solvents, water content (if reported), and storage recommendations. Align method validation (selectivity, carryover, stability) with batch data.

Reaction and Applications

Primary application: stable‑isotope internal standard for LC–MS/MS quantitation of rifampicin in research matrices and for method development/validation. Deuteration at eight positions yields a defined mass shift with closely matched chromatographic behavior.

• Quantitative LC–MS/MS (general guidance):

  • Use rifampicin‑d8 as an internal standard for calibration curves (matrix‑matched), recovery studies, and stability testing (bench‑top, freeze–thaw, autosampler).
  • MS settings (example, literature): ESI+ with multiple reaction monitoring (MRM) targeting parent m/z [M+H]+ shifted by +8; fragment transitions selected to avoid back‑exchange or H/D scrambling.

• Sample types (research only):

  • Biological matrices (e.g., preclinical plasma/serum, tissue homogenates), environmental samples, and formulation extracts. Spike IS prior to extraction to correct for recovery and matrix effects.

• Chemistry/derivatization:

  • Generally analyzed underivatized. For special workflows, mild derivatization of phenolic groups is possible (e.g., silylation for GC–MS) but less common due to complexity.

• Practical tips:

  • Protect from light during all prep and autosampler storage.
  • Control pH of extraction buffers (avoid strong base/acid) to minimize hydrolysis/oxidation. Include antioxidants (ascorbate) only if validated not to interfere.
  • Evaluate isotopic purity and potential co‑elution of partially deuterated species; set MRM windows accordingly.

• Method validation checkpoints (ICH/US‑FDA bioanalytical guidance—general): selectivity, calibration curve model, accuracy/precision, recovery, matrix factor, carryover, dilution integrity, stability (short‑term, long‑term, freeze–thaw, post‑prep). Note: applicable to research method development; adapt to institutional standards.

Reaction Conditions

As an analytical internal standard, rifampicin‑d8 is not typically subjected to preparative reactions. The following guidance focuses on conditions relevant to extraction, chromatographic separation, and stability in research workflows (literature/general):

• Extraction/sample prep:

  • Protein precipitation: add 3–5 volumes of cold ACN or MeOH containing rifampicin‑d8 (IS) to biological matrix; vortex, centrifuge, and collect supernatant.
  • SPE: weak cation exchange (due to piperazine) or reversed‑phase sorbents; condition with MeOH and water, load sample with IS, wash with aqueous buffer, elute with MeOH/ACN containing 0.1–1% NH4OH or formic acid depending on mode.
  • LLE: ethyl acetate or MTBE after pH adjustment; less preferred environmentally than SPE/PP.

• Chromatography:

  • LC columns: C18 or polar‑embedded phases; consider phenyl‑hexyl for improved retention/selectivity.
  • Mobile phases: water (0.1% formic acid or 2–10 mM ammonium formate/acetate) and ACN/MeOH. Gradient elution typical. Protect autosampler vials from light (amber glass) and keep at 4–10°C during runs.

• Mass spectrometry:

  • Ionization: ESI positive. Monitor [M+H]+ of analyte and +8 Da for IS. Choose product ions that retain deuterium labels to avoid overlap.

• Stability windows (literature, parent):

  • Bench‑top: hours in subdued light at neutral pH; degradation accelerates under bright light and extreme pH.
  • Autosampler: stable up to 24–48 h at 4–10°C protected from light; confirm by stability experiments.

All parameters should be verified and optimized per method‑development best practices; values above are general literature‑based guidance, not specifications.

Safety and Handling

Authoritative safety data for this specific item (GHS signal word, H‑statements, pictograms) are Not specified for this item; refer to CoA/Spec Sheet and the product SDS. The following are general laboratory safety considerations for rifamycin‑class compounds and isotope‑labeled standards.

• General hazards (literature/general):

  • May cause skin/eye irritation; harmful if swallowed or inhaled. Fine powders can be dust‑forming—handle in a fume hood or containment.
  • Rifampicin is photosensitive; light can induce degradation—protect from light to avoid formation of irritant/decomposition products.
  • Not for human or veterinary use. For research use only.

• PPE: lab coat, safety glasses or face shield, and appropriate chemically resistant gloves (e.g., nitrile). Use a dust mask/respirator if aerosolization risk; otherwise work in a fume hood.

• Handling practices:

  • Minimize exposure to light and air; keep containers tightly closed. Use amber vials for solutions.
  • Avoid contact with strong oxidizers and strong acids/bases that can accelerate degradation (literature guidance).
  • Prevent cross‑contamination of bioanalytical labs; isotopically labeled internal standards can confound LC–MS assays if spilled.

• First aid (general):

  • Skin/eye contact: rinse with water for 15 minutes; remove contaminated clothing; seek medical advice.
  • Inhalation: move to fresh air; seek medical attention if symptoms persist.
  • Ingestion: rinse mouth; seek medical attention.

• Waste: Collect solutions/solids in halogen‑free organic waste appropriate for antibiotics/organics. Follow institutional/EHS disposal procedures.

Always consult the SDS for definitive hazard classification and emergency measures.

Solvent Selection

Rifampicin‑d8 behaves like rifampicin in solvent affinity; deuteration minimally affects solubility.

• Polarity/miscibility (literature/general):

  • Highly soluble in DMSO and DMF; good in methanol and acetonitrile; limited in water at neutral pH due to amphoteric aggregation.
  • A small fraction of acid or base (e.g., 0.1% formic acid or 5–10 mM ammonium bicarbonate) can enhance solubility in aqueous-organic mobile phases for LC–MS.

• Recommended stock solvents (analytical use):

  • DMSO (preferred for long‑term concentrated stocks; excellent chemical stability and minimized evaporation).
  • Methanol or acetonitrile (for immediate use working solutions; rapid mixing with mobile phase).

• When to choose alternatives:

  • Use methanol/acetonitrile when DMSO is incompatible with bioassays or when viscosity must be minimized.
  • Avoid chlorinated solvents and strong protic acids/bases that may promote degradation or salt exchange.

• Small comparison (literature trends):

  • DMSO: highest solubility/stability; viscous; can suppress ESI if injected at high %.
  • MeOH: good solubility; enhances ESI; may increase degradation on prolonged storage.
  • ACN: moderate solubility; excellent for LC–MS; lower proticity may reduce hydrolysis.

Practical tip: Prepare a 1–10 mg/mL stock in DMSO (amber vial), then dilute 1:100–1:1000 into initial mobile phase (e.g., water/ACN with 0.1% formic acid or ammonium formate) immediately before calibration/quantitation. Filter only if necessary (PTFE, low‑adsorption) to avoid analyte loss.

Storage and Reconstitution

Item-specific storage from Product Data:

• Storage Conditions: Room temperature.
• Shipped In: Not specified for this item; refer to CoA/Spec Sheet.

General best practices (literature/general; complement, not override, the above):

• Solid material:

  • Store tightly closed at room temperature in a dry, dark place (use amber glass; protect from light). Consider desiccation and inert gas backfill to limit oxidation.
  • Avoid humid environments that can induce clumping or gradual degradation.

• Reconstitution:

  • Prepare concentrated stock solutions in anhydrous DMSO (e.g., 1–10 mg/mL). Mix gently to dissolve fully; brief sonication can help.
  • For immediate use, dilute the DMSO stock into MeOH or ACN, then into aqueous mobile phase (e.g., with 0.1% formic acid or 2–10 mM ammonium formate). Prepare fresh working solutions daily when possible.

• Solution storage (general guidance):

  • Aliquot stocks into amber vials to minimize headspace and freeze–thaw cycles. Although the solid is stored at room temperature per product data, refrigerating or freezing solutions (e.g., −20°C) can improve stability; validate stability under your conditions.
  • Protect all solutions from light. Limit autosampler residence time; cool autosampler (4–10°C) when available.

• Stability monitoring:

  • Verify short‑term and long‑term solution stability, freeze–thaw robustness, and post‑prep stability during method development. Replace solutions showing precipitate, color change, or unexpected LC–MS responses.

Always consult the product CoA/SDS for lot‑specific guidance.

Structure and Identity

Brief: Rifampicin‑d8 is the stable‑isotope–labeled analogue of rifampicin with eight non‑exchangeable deuterium atoms, designed for quantitative LC–MS/MS as an internal standard.

• Item-specific identifiers (this product):

  • CAS: Not specified for this item; refer to CoA/Spec Sheet.
  • InChIKey: Not specified for this item; refer to CoA/Spec Sheet.
  • SMILES: 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.

• Literature/typical identity information (for context):

  • Parent rifampicin (non‑deuterated) formula (literature): C43H58N4O12; average MW ≈ 822.94 g/mol.
  • Theoretical average MW for d8 isotopologue (computed): ≈ 822.94 + 8.06 ≈ 831.0 g/mol (exact value depends on labeling pattern).
  • Compound class: ansamycin (rifamycin family) bearing a naphthohydroquinone chromophore bridged by an aliphatic “ansa” chain and a substituted hydrazone linking to a 4‑methylpiperazine moiety.

• Structural features (descriptive, literature):

  • Aromatic naphthoquinone/hydroquinone core with multiple phenolic –OH groups.
  • Long aliphatic ansa bridge decorated with secondary/tertiary alcohols and an amide/hydrazone functionality.
  • Terminal tertiary diamine (4‑methylpiperazine) confers basicity and amphiphilicity.
  • Multiple H‑bond donors/acceptors; high polarity yet overall lipophilic surface.
  • Rifampicin‑d8 retains the parent topology; eight hydrogens on defined carbon positions are replaced by deuterium (precise labeling pattern may vary by supplier; consult CoA for positional labeling).

• 2D structural description in words (literature): A naphthalene‑derived quinone ring substituted by phenolic groups, connected via an aliphatic macrocyclic “ansa” chain bearing several hydroxyls and carbonyls to an imino/hydrazone linkage terminating in a 4‑methylpiperazine ring; stereocenters reside along the ansa chain.

Synthetic Utility

Rifampicin‑d8 is not commonly used as a synthetic building block due to its structural complexity and cost; its primary value is analytical. However, from a synthetic and mechanistic perspective, it offers unique utilities:

• Isotope tracing (general):

  • Enables tracking of degradation pathways (photooxidation, hydrolysis, imine/hydrazone exchange) by monitoring retention of deuterium labels in fragments, aiding in mechanism elucidation.

• Method development tool:

  • Facilitates optimization of sample‑prep recovery (protein precipitation, SPE, LLE) by allowing direct mass‑balanced comparisons of IS and analyte.

• Reactivity of functional groups (literature, parent rifampicin):

  • Phenolic groups can undergo acylation/alkylation under mild conditions.
  • Hydrazone/imine linkage is susceptible to hydrolytic cleavage and transimination under acidic or basic conditions.
  • Secondary/tertiary alcohols along the ansa chain can form esters/ethers; oxidation can occur at hydroquinone sites.

• Retrosynthetic value (academic interest):

  • The ansamycin framework illustrates macrocyclization strategies and late‑stage diversification on complex polyketide‑derived scaffolds. Deuterium incorporation patterns reflect H/D exchange or incorporation steps in total synthesis/semisynthesis studies.

In practice, reserve rifampicin‑d8 for analytical roles; employ unlabeled rifampicin or model fragments for exploratory reactivity studies to conserve labeled material.

Target Specificity

Not applicable. This product is a small‑molecule isotope‑labeled standard, not an antibody, enzyme, nucleic acid, or affinity reagent. There are no clone/isotype, antigen, or species‑reactivity attributes. For biological interaction context, see the “Biological Roles” section.