solutions for oligonucleotides synthesis liquid reagents from safc supply solutions®

ProClin® Preservatives Mechanisms and Stability

for Diagnostics Reagents

www.safcsupplysolutions.com

Solutions for Oligonucleotides SynthesisLiquid Reagents

from SAFC Supply Solutions®


www.sigma-aldrich.com


2

SAFC Supply Solutions® has more than 25 years of manufacturing experience in DNA and RNA synthesis reagents. Our liquid reagents meet the most stringent quality specifications, making them ideal for use in automated nucleic acid synthesis. Lot-to-lot consistency is guaranteed for every product manufactured at SAFC Supply Solutions ISO 9001 certified site and each lot is quality-controlled to ensure optimum synthesis performance.

Chemical composition and purpose

Deblock solution containing trichloroacetic acid (TCA) or dichloroacetic acid (DCA) in dichloromethane or toluene removes the dimethoxytrityl (DMT) protecting group from the 5’hydroxyl moiety of nucleotides

already incorporated into the growing nucleic acid, prior to the addition of the next phosphoramidite. Removal of the DMT allows the unprotected 5’hydroxyl moiety to react with a new phosphoramidite in a subsequent extension reaction.

Wash solution containing anhydrous acetonitrile (water content ≤30 ppm), is used as the overall wash solvent following each step in the synthesis cycle.

Activator solution containing powerful activators like 4,5-dicyanoimidazole (DCI) or 5-(3,5-bis(trifluoromethyl) phenyl)-1H-tetrazole (Activator 42) in acetonitrile are mixed with solutions of phosphoramidites during the extension step. The activator reacts with the amidite group to form a highly reactive intermediate. The intermediate forms an internucleotide bond with the detritylated 5’hydroxyl group of the growing oligonucleotide chain.

Cap A solution containing acetic anhydride and tetrahydrofurane (THF) is used after the phosphoramidite reaction/coupling step. This solution aborts chains bearing unreacted 5’hydroxyl groups due to failure of reaction with activated phosphoramidite (typically, 1 to 2% of growing oligonucleotide chains in each synthesis cycle). These unreacted 5’hydroxyl groups are capped with an acetyl group, thus rendered unreactive for subsequent synthesis steps. For synthesis with Proligo® Reagents’ TAC-protected phosphoramidites, Fast Deprotection CAP A solution must be employed instead of Cap A solution.

Purity of Liquid Reagents to support your overall synthesis performanceIn solid-phase synthesis of oligonucleotides, liquid reagents are used in each

step of the synthesis cycle. Overall synthesis performance, and therefore total

product yield and purity of the crude oligonucleotide, is highly dependent on

the chemical purity of the monomers and the supporting liquid reagents.

Features Benefits

Products designed for oligonucleotide synthesis

Constant high synthesis performance

Liquid reagents offered in addition to amidites and supports

Key oligonucleotide synthesis reagents available in one shop

Constant high control

Matching with high quality amidites and support products

Broad range of packaging

Offering high flexibility

Customized formulations and specifications

Special requirements will be fulfilled

Technical expertise Supporting customized solutions

New product development: Activator® 42

Synthesis optimization and cost reduction

Table 1. Liquid Reagent features and benefits




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Fast Deprotection Cap A solution must be used for synthesis with Proligo Reagents’ TAC-protected phosphoramidites. Fast Deprotection Cap A solution, with tert-butylphenoxyacetyl acetic anhydride (TAC2O) in tetrahydrofuran is used in place of Cap A solution to ensure that the displacement of tert-butylphenoxyacetyl (TAC) on guanine bases of the TAC-protected phosphoramidites does not occur.

Cap B solution containing tetrahydrofurane (THF) pyridine and N-methylimidazole (NMI),

is mixed in situ with Cap A solution during the capping (acetylation) reaction. Pyridine is applied as a mild base while NMI provides a powerful acylation catalyst.

Oxidizer solution, containing tetrahydrofurane (THF), pyridine, iodine and water is applied to oxidize the chemically unstable trivalent phosphorous triester linkage formed in the coupling reaction to a stable pentavalent phosphate triester. Iodine functions as a mild oxidant while water serves as an oxygen donor.

The Oligonucleotide Synthesis Cycle



4

A broad range of packaging

Flexible packaging options are available to meet most synthesizer requirements, including ABI® 394, ABI® 3900, ExpediteTM, ÄKTATM and Mermade. As a single source supplier, SAFC Supply Solutions’ Hamburg (Germany) and Sheboygan, Wisconsin manufacturing sites can liquid fill and blend material in and have complete packaging

operations that can accommodate small-scale bottles to 1,400 L returnable containers.

Our worldwide distribution network offers advantages of dedicated airfreight space, expertise in chemical transport regulations and local delivery knowledge.

BottlesDescription Height Diameter Capacity Total Capacity Material Closure

100 mL Septum 95 mm 51 mm 100 mL 120 mL amber glass septum

4 oz 22-400 Bottle 112 mm 48,5 mm 120 mL 125 mL amber glass screw cap 22-400

8 oz 24-400 Bottle 137 mm 60 mm 200 mL 240 mL amber glass screw cap 24-400




2 L 38-430 Bottle 310 mm 135 mm 2 L 2.5 L amber glass screw cap 38-430

2.5 L GL 45 Bottle 296 mm 135 mm 2.5 L 2750 mL amber glass screw cap GL 45

4 L 38-430 Bottle 353 mm 161 mm 4 L 4300 mL amber glass screw cap 38-430

1 L GL 45 Bottle 212 mm 101 mm 1000 mL 1158 mL amber glass screw cap GL 45

100 mL Septum 4 oz 22-400 Bottle 8 oz 24-400 Bottle 8 oz 28-400 Bottle 16 oz 28-400 Bottle

32 oz 28-400 Bottle 2 L 38-430 Bottle 2.5 L GL 45 Bottle 4 L 38-430 Bottle 1 L GL 45 Bottle

Table 2. Worldwide Packaging Options




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Drums

Description Capacity Total Capacity Weight Height Diameter Wall

Thickness Material Closure Operating Pressure

Approval Number

18 L Returnable Drum

Stainless steel drum 18 L with Dip Tube

18 L 20 L 6,6 kg 442 mm 278 mm 1.5 mm V2A 1.4301 SS304

Screw cap DIN 11851 PTFE R52x1/6”

0.5 bar 1A1/X2,0/900

25 L Drum, Disposal

Composite drum sheet steel/PE

25 L 30 L 3.4 kg 560 mm 280 mm 0.5 mm PE-HD A BCS 56x4 (1x) 1A1/X1,6/250


Stainless Steel Drum with Dip Tube

25 L 30 L 10 kg 400 mm 380 mm 1.5 1.4301 3A Coding MAV Micro Matic Valve

0.5 bar 1A1/X2,3/900



45 L 50 L 11 kg 600 mm 363 mm 1.5 mm V2A 1.4301 SS304

Screw cap DIN 11851 PTFE R52x1/6”

0.5 bar 1A1/X2,3/900


Stainless Steel Drum with Dip Tube

45 L 50 L 12 kg 600 mm 263 mm 1.5 mm 1.4301 4F Coding MAV Micro Matic Valve

0.5 bar 1A1/X2,3/900

180 L Drum, Disposal

Composite drum sheet steel/PE

180 L 200 L 21 kg 884 mm 585 mm 1 mm PE-HD BCS 56x4 (2x) 1A1/X1,3/250

200 L Returnable Drum, Level Sensor

Stainless steel drum 200 L with level sensor and Dip Tube

200 L 225 L 43 kg 970 mm 600 mm 2 mm 1.4301 On Demand 0.5 bar 1A1W/X2,0/900

200 L Returnable Drum, Standard


200 L 225 L 43 kg 970 mm 600 mm 2 mm 1.4301 Screw cap DIN 11851 PTFE R52x1/6”

0.5 bar 1A1W/X2,0/900

Returnable 1400 L Container

Stainless Steel IBC 1400 L with Dip Tube

1400 L 1500 L 247 kg 1960 mm 1200 mm 1.4301 On demand 0.5 bar 31A/Y

18 L Returnable Drum 25 L Drum, Disposal 25 L Returnable Drum 45 L Returnable Drum 45 L Returnable Drum

180 L Drum, Disposal 200 L Returnable Drum, Level Sensor

200 L Returnable Drum, Standard

Returnable 1400 L Container

Table 3. Liquid Reagent Packaging



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For special requirements on formulations,

specifications and packaging, our manufact-

uring sites offer custom blending services:

• Batch sizes from 100 L up to 4000 L

• A wide variety of high purity solvents, including flammable solvents handling

• Raw materials supplier qualification process in alignment with customer expectations

• Specifications and Quality Control according to customer requirements

Six additional production plants support the Sheboygan and Hamburg facilities with custom blending and risk mitigation for all projects.

All inquiries are evaluated by a Project Management Group that selects the most appropriate manufacturing site and follows up on customer projects.

Custom Blending Solutions to speed up your processes

Description Capacity Total Capacity Weight Height Diameter Wall

Thickness Material Closure Operating Pressure

Approval Number

20 L Pure-Pac II 20 L 22 L 8 kg 20 inches 11 inches 1.5 mm SS304 MAV Micro Matic Valve - Simriz 485

0.34 bar 1A1/X2.3/900

50 L Pure-Pac II 50 L 55 L 15 kg 23 inches 16 inches 1.5 mm SS304 MAV Micro Matic Valve - Simriz 485

0.34 bar 1A1/X2.0/600

200 L Pure-Pac II 200 L 220 L 47 kg 38 inches 24 inches 2 mm SS304 MAV Micro Matic Valve - Simriz 485

0.34 bar 1A1/X2.0/600

200 L Pure-Pac II, level sensor

200 L 220 L 47 kg 38 inches 24 inches 2 mm SS304 MAV Micro Matic Valve - Simriz 485

0.34 bar 1A1/X2.0/600

Additional Drum Options (US market only)

For your custom packaging requirements, including technical and product specifications, please contact your sales representative to inquire. Some packaging options may not be available in your region.

Table 4. Liquid Reagent Packaging




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7

Reagent Description Packaging Available

Formulation Specifications Product Number Unit Bottle Type

Amidite Diluent & Acetonitrile

ACN Water Content ≤ 30ppm or ≤10ppm also available *

L010010-06 6 x 100 mL 100 mL septum

nD20 = 1,342-1,346 L010200-04 4 x 2 L 2 L 38/430

App. of Sol. = clear L010250-04 4 x 2.5 L 2.5 L GL45

L010400-04 4 x 4 L 4 L 38/430

L011800-01 1 x 18 L 18 L Returnable Drum

L014500-01 1 x 45 L 45 L Returnable Drum

L014500-C01-01 1 x 45 L 45 L Returnable Drum

L0120000-01 1 x 200 L 200 L Returnable Drum 2” Level Sensor

L0120000-C01-01 1 x 200 L 200L Returnable Dru m 2” Standard

L0114000-01* 1 x 1400 L Returnable 1400L Container

L810045-06 6 x 450 mL 16 oz 28/400 (450 mL) for PE 8900/Polygen

TFA Deblock Toluene Toluene/TFA Water Content ≤ 200 ppm

L0202502-04 4 x 2.5 L 2.5 L GL45

100/1 TFA Content = 10.3-12.5 g/l

v/w App. of Sol. = clear

DCA Deblock DCM/DCA Water Content ≤ 200 ppm

L020251-04 4 x 2.5 L 2.5 L GL45

97/3 DCA Content = 42.2-51.6 g/l

v/v App. of Sol. = clear

TCA Deblock DCM/TCA Water Content ≤ 200 ppm

L020250-04 4 x 2.5 L 2,5 L GL45

100/3 TCA Content = 28-32 g/l

L020400-04 4 x 4 L 4 L 38/430

v/w App. of Sol. = clear L022500-01 1 x 25 L 25 L Drum, Disposal

L320045-01 1 x 450 mL 16 oz 28/400 (450 mL)


Activator 42 0.1M ACN/Activator42 Water Content ≤ 30ppm

L0302502-04 4 x 2.5 L 2,5 L GL45

100/3 Act. 42 Content = 28-31 g/l

L3300182-06 6 x 180 mL 8 oz 24/400 (200 mL) for ABI

v/w App. of Sol. = clear L8300202-06 6 x 200 mL 8 oz 28/400 (200 mL) for PE 8900/Polygen

L8300452-06 6 x 450 mL 16 oz 28/400 (450 mL)

Table 5. Liquid Reagent Formulations



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Activator 42 0.25 M ACN/Activator42 Water Content ≤ 30 ppm

L0302512-04 4 x 2.5 L 2.5 L GL45

100/7 Act. 42 Content = 70-77 g/l


v/w App. of Sol. = clear L8300462-06 6 x 450 mL 16 oz 28/400 (450 mL)

ETT Activator 0.25M ACN/ETT Water Content ≤ 30 ppm

L0302501-04 4 x 2.5 L 2.5 L GL45

100/3 ETT Content = 32-35 g/l

L0302001-04 4 x 2 L 2 L 38/430

v/w App. of Sol. = clear L8300201-06 6 x 200 mL 8 oz 28/400 (200 mL) for PE 8900/Polygen

L8300451-06 6 x 450 mL 16 oz 28/400 (450 mL)

DCI Activator 0.25M ACN/DCI Water Content ≤ 30 ppm

L080250-04 4 x 2.5 L 2,5 L GL45

100/3 DCI Content=29-32 g/l L080400-04 4 x 4 L 4 L 38/430

v/w App. of Sol. = clear L082500-01 1 x 25 L 25 L Returnable Drum

L380018-06 6 x 180 mL 8 oz 24/400 (200 mL) for ABI

L880020-06 6 x 200 mL 8 oz 28/400 (200 mL)

L880045-06 6 x 450 mL 16 oz 28/400 (450 mL)

Cap A THF/Ac2O Ac2O Content = 88.7-108.4 g/l

L040250-01 1 x 2.5 L 2.5 L GL45

90.9/9.1 App. of Sol. = clear L040250-04 4 x 2.5 L 2.5 L GL45

v/v L040400-01 1 x 4 L 4 L 38/430

L040400-04 4 x 4 L 4 L 38/430

L040200-04 4 x 2 L 2 L 38/430


L840045-06 6 x 450 mL 16 oz 28/400 (450 mL)

Cap A for ABI instruments

THF/Lutidine/Ac2O Ac2O Content = 97.0-119.0 g/l

L040251-01 1 x 2.5 L 2.5 L GL45

80/10/10 App. of Sol. = clear, colorless to brownish solution

L340018-06 6 x 180 mL 8 oz 24/400 (200 mL)

v/v/v L340045-06 6 x 450 mL 16 oz 28/400 (450 mL)

Cap B THF/Py/NMI Water Content ≤ 200 ppm

L050250-01 1 x 2.5 L 2.5 L GL45

80/10/10 Formulation (GC) conforms

L050250-04 4 x 2.5 L 2.5 L GL45

v/v/v App. of Sol. = clear L050400-01 1 x 4 L 4 L 38/430

L050400-04 4 x 4 L 4 L 38/430

L050200-04 4 x 2 L 2 L 38/430







9



Cap B for ABI instruments

THF/NMI Water Content ≤ 200 ppm

L050251-01 1 x 2.5 L 2.5 L GL45

84/16 Formulation (GC) conforms

L350018-06 6 x 180 mL 8 oz 24/400 (200 mL)

v/v App. of Sol. = clear L350045-06 6 x 450 mL 16 oz 28/400 (450 mL)

Fast Deprotection Cap A THF/Tac2O Tac2O Content = 45-55 g/l

L070250-01 1 x 2.5 L 2.5 L GL45

100/5 App. of Sol. = clear L070400-01 1 x 4 L 4 L 38/430

v/w L370018-06 6 x 180 mL 8 oz 24/400 (200 mL) for ABI

L370045-06 6 x 450 mL 16 oz 28/400 (450 mL)

L870020-06 6 x 200 mL 8 oz 28/400 (200 mL) PE 8900/Polygen

Oxidizer 0.02M THF/H2O/Py/Iodine Iodine Content = 3.9-4.7 g/l

L060250-04 4 x 2.5 L 2.5 L GL45

90.54/9.05/0.41/0.43 App. of Sol. = brownish L060400-04 4 x 4 L 4 L 38/430

v/v/v/w L060200-04 4 x 2 L 2 L 38/430

L860021-06 6 x 200 mL 8 oz 24/400 (200 mL) for ABI



Oxidizer 0.1M for ABI instruments

THF/H2O/Py/Iodine Iodine Content = 22.9-27.9 g/l

L060251-04 4 x 2.5 L 2.5L GL45

77/2/21/2.54 App. of Sol. = brownish

L360020-06 6 x 200 mL 8 oz 24/400 (200 mL)

v/v/v/w L360045-06 6 x 450 mL 16 oz 28/400 (450 mL)

Oxidizer 0.02M for ABI 3900 instruments

THF/H2O/Py/Iodine Iodine Content = 5.4-6.6 g/l

L060252-04 4 x 2.5 L 2.5 L GL45

66/12/22/0.6 App. of Sol. = brownish

L260045-06 6 x 450 mL 16 oz 28/400 (450 mL)

v/v/v/w

ACN Based Liquid Reagents

Cap A ACN ACN/Ac2O Ac2O Content = 88.7-108.4 g/l

LB40250-01 1 x 2.5 L 2.5 L GL45

90.9/9.1 App. of Sol. = clear

v/v

Cap B ACN ACN/Py/NMI Water Content ≤ 100 ppm

LB50250-01 1 x 2.5 L 2.5 L GL45


v/v/v App. of Sol. = clear

Cap 1 in ACN ACN/Lutidine/NMI Water Content ≤ 200 ppm

LR40250-01 1 x 2.5 L 2.5 L GL45


v/v/v App. of Sol. = clear




10



Fast Deprotection Cap 2 in ACN

Tac2O/ACN Tac2O Content = 45-55g/l

LR50250-01 1 x 2.5 L 2.5 L GL45

100/5 App. of Sol. = clear

v/w

Oxidizer in ACN ACN/H2O/Py/Iodine Iodine Content = 3.9-4.7 g/l

LR60250-04 4 x 2.5 L 2.5 L GL45

90.54/9.05/0.41/0.43

v/v/v/w

ÄKTA Solution

DCA Deblock (Toluene) DCA/Toluene Water Content ≤ 200 ppm

L520251-04 4 x 2.5 L 2.5 L GL45

97/3 DCA Content = 42.2-51.6


BTT Activator 0.3 M 2.5 L, ÄKTA

ACN/BTT Water Content ≤ 30 ppm

L580250-04 4 x 2.5 L 2.5 L GL45

100/6 BTT Content = 51.9-63.4

v/w App. of Sol. = clear

Cap A for Äkta and Oligo Pilot

ACN/NMI Water Content ≤ 100 ppm

L540250-01 1 x 2.5 L 2.5 L GL45



Cap B1 for Äkta and Oligo Pilot

ACN/Ac2O Formulation (GC) conforms

L550250-01 1 x 1.25 L 2.5 L GL45

60/40 App. of Sol. = clear

v/v

Cap B2 for Äkta and Oligo Pilot

ACN/Py Water Content ≤ 200 ppm

L550251-01 1 x 1.25 L 2.5 L GL45



Oxidizer 0,05M for Äkta and Oligo Pilot

Iodine/Py/H2O Iodine Content = 11.4-14.0 g/l

L560250-04 4 x 2.5 L 2.5 L GL45

0,05M/90/10 App. of Sol. = clear

m/v/v





11

Quality Control

SAFC has rigorous QC protocol for every product produced to identify purity levels against set specifications, including custom QC testing. State-of-the-art analytical methods include:

• RP & SAX HPLC

• LC-MS

• NMR (31P, 1H, 13C, 19F)

• Nucleic acid synthesis test

• FT-IR

• Titration & “Wet Chemistry”

• UV/VIS

• Karl Fischer water analysis

• Mercury porosimetry

Quality Assurance

As a trusted partner to the oligonucleotide market, SAFC assists its customers in new product development. We know that developing materials includes having insight to their quality requirements. As a result, our company culture fosters continuous process improvement and employs quality systems to provide complete traceability on all our raw materials, processes and procedures. We serve customers conducting audits with complete documentation and files; our quality assurance department routinely accepts requests for documentation, including Certificates of Origin. Our Hamburg facility was one of the first in the industry to be ISO 9001 certified (1993), and we have synthesized amidites and liquid reagents under ISO9000:2000 since then.

Abbreviations

ACN Acetonitrile

DCM Dichloromethane

TCA Trichloroacetic Acid

TFA Trifluoroacetic Acid

DCA Dichloroacetic Acid

THF Tetrahydrofurane

Ac2O Acetic Acid Anhydride

Py Pyridine

NMI N-Methylimidazole

Tac2O tert. Butylphenoxyacetyl Acetic Acid Anhydride

DCI 4,5-Dicyanoimidazole

ETT 5-Ethylthio-1H-tetrazole


KUC04695-5031200049

SAFC®, SAFC Supply Solutions® and Sigma-Aldrich® are registered trademarks of Sigma-Aldrich Biotechnology L.P. and Sigma-Aldrich Co.

Proligo® Reagents is a registered trademark of Sigma-Aldrich Biotechnology L.P. and Sigma-Aldrich Co. Activator® 42 is a registered trademark of Sigma-Aldrich

Biotechnology L.P. and Sigma-Aldrich Co. ABI® is a registered trademark of Applera Corporation or its subsidiaries. ÄKTA™ oligopilot is a trademark of GE

Healthcare. Expedite™ is a trademark of Applera Corporation or its subsidiaries.

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