Protein and peptide drugs hold great promise as therapeutic agents. However, many are degraded by proteolytic enzymes, can be rapidly cleared by the kidneys, generate neutralizing antibodies and have a short circulating half-life. Pegylation, the process by which polyethylene glycol chains are attached to protein and peptide drugs, can overcome these and other shortcomings. By increasing the molecular mass of proteins and peptides and shielding them from proteolytic enzymes, pegylation improves pharmacokinetics. This article will review how PEGylation can result in drugs that are often more effective and safer, and which show improved patient convenience and compliance.

[1]. Adsorption of polyamine, polyacrylic acid and polyethylene glycol on montmorillonite: An in situ study using ATR-FTIR. Volume 14, Issue 1, March 1997, Pages 19-34.

[2]. Structural basis of polyethylene glycol recognition by antibody. J Biomed Sci. 2020 Jan 7;27(1):12.

[3]. Effect of pegylation on pharmaceuticals. Nat Rev Drug Discov. 2003 Mar;2(3):214-21.

[4]. Injectable silk-polyethylene glycol hydrogels. Acta Biomater. 2015 Jan;12:51-61.

[5]. Beneficial effects of combining nilotinib and imatinib in preclinical models of BCR-ABL+ leukemias. Blood. 2007 Mar 1;109(5):2112-20.

Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray diffraction (XRD) were used to investigate the adsorption process of water-soluble polymers from aqueous solutions onto the surface of clay minerals in situ, aiming to understand the microscopic interactions in water-based mud (WBM). The water-soluble polymers studied included polyethylene glycol (PEG) (molecular weights Mw 300 and 1000, respectively) (neutral), polyamine (FL15) (Mw 5000) (cationic), and polyacrylic acid (PAA) (Mw 2000) (charge dependent on pH), with the aim of determining the effect of polymer charge on the adsorption properties and extent. XRD and ATR-FTIR spectroscopy results showed that the water-soluble polymers could adsorb onto the clay dispersion and remain stable when deposited into thin solid films. XRD results indicated that PEG was stacked in monolayer or bilayer form, while PAA and FL15 were confined to monolayers between clay layers. ATR-FTIR spectroscopy showed that FL15 could penetrate into Na-SWy-1 films with low PEG loading without displacing any of the original PEG. FL15 could not penetrate the film when bilayer PEG was present. Both PAA and PEG were adsorbed onto the Na-SWy-1 film loaded with FL15, and the amount of adsorption was independent of the loading of FL15 or the molecular weight of PEG. ATR-FTIR results showed that significant adsorption occurred within 30 seconds and the adsorption rate was not affected by the second polymer preloaded into the clay. [1]

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Background: Polyethylene glycol (PEG) is widely used in industrial and pharmaceutical fields. Anti-PEG antibodies have been developed for characterizing PEGylated drugs and other applications. However, the potential mechanism of specific PEG binding has not been elucidated. Methods: Fab fragments of two homologous anti-PEG antibodies, 3.3 and 2B5, were complexed with PEG and crystallized, and their structures were determined by X-ray diffraction. The PEG-Fab interaction in the two crystals was analyzed and compared with the interaction in the PEG-containing crystal of the unrelated antihemagglutinin 32D6-Fab. The stoichiometry of PEG binding was determined by analytical ultracentrifugation (AUC). Results: The PEG binding modes of 3.3 and 2B5 were similar, both exhibiting an S-shaped core PEG fragment binding to two dimer-associated Fab molecules. Nearby satellite binding sites may accommodate portions of longer PEG molecules. The core PEG fragment primarily interacted with heavy chain residues D31, W33, L102, Y103, and Y104, and formed extensive contacts with aromatic side chains. At the center of each semicircle of the S-shaped PEG, a water molecule alternately formed hydrogen bonds with ether oxygen atoms, with a configuration similar to crown ether-bound lysine. Each satellite fragment was sandwiched between two arginine residues (R52 on the heavy chain and R29 on the light chain) and also interacted with multiple aromatic side chains. In contrast, the non-specifically bound PEG fragments in the 32D6-Fab crystal were located in the elbow region or lattice contacts. AUC data indicated that 3.3-Fab existed as a monomer in PEG-free solutions but formed a dimer in the presence of PEG-550-MME, with the dimer size similar to the S-shaped core PEG fragment. Conclusion: Different amino acids in 3.3 and 2B5 do not participate in PEG binding, but participate in dimer formation. Specifically, the light chain residue K53 of 2B5-Fab has significant contact with another Fab in the dimer, while the corresponding residue N53 of 3.3-Fab does not. This difference in protein-protein interaction between the two Fab molecules in the dimer may explain the temperature dependence of PEG binding of 2B5 and the inhibitory effect of crown ether. [2]

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Silk fibroin hydrogels for tissue repair are usually prepared in advance by chemical or physical methods. For many medical applications, it is ideal to use high concentrations (>8%) of injectable silk fibroin hydrogels to avoid surgical implantation and achieve slow degradation of the gel in vivo. In this study, injectable silk fibroin solutions that can form hydrogels in vivo were prepared by mixing silk fibroin with low molecular weight polyethylene glycol (PEG), particularly PEG300 and 400, with molecular weights of 300 and 400 g mol⁻¹, respectively. The gelation time depends on the concentration and molecular weight of PEG. When the PEG concentration in the gel reaches 40-45%, the gelation time is less than 30 minutes, which is confirmed by optical density measurements and rheological studies. Furthermore, the gelation kinetics of PEG400 are faster than those of PEG300. Gelation is accompanied by structural changes in silk fibroin, leading to the transformation of random coils in solution into crystalline β-sheet structures in the gel, which is confirmed by circular dichroism spectroscopy, attenuated total reflectance Fourier transform infrared spectroscopy, and X-ray diffraction. The measured modulus (127.5 kPa) and yield strength (11.5 kPa) are comparable to those of ultrasound-induced hydrogels at the same silk fibroin concentration. The injection performance of 15% PEG-silk fibroin hydrogel injected through a 27G needle changes over time; the compressive force gradually increases from approximately 10 N to 50 N within 60 minutes. The growth of human bone marrow mesenchymal stem cells on PEG-silk fibroin hydrogel is inhibited, possibly due to the presence of PEG. Cell growth only begins after 5 days, presumably due to the dissolution of PEG from the gel. When 5% PEG-silk fibroin hydrogel was subcutaneously injected into rats, ultrasound imaging and histological analysis showed that the hydrogel was significantly degraded and tissue ingrowth occurred after 20 days. No obvious inflammatory response was observed around the gel. These hydrogels are characterized by injectability, slow degradation and low initial cell adhesion, indicating their potential value in many biomedical applications, such as antifouling and antiadhesion. [4]

(C2H4O)NH2O

35000

25322-68-3

White to off-white solid powder

1.125

250ºC

-65ºC

171ºC

<0.01 mm Hg ( 20 °C)

1.458-1.461

0

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)