Table 3

Purification of insulin precursor by IMAC and conversion to human insulin

Steps

Insulin species

(molecular mass)

Amount (mg) ^a

Amount (mmol) ^a

Recovery (%) ^a

Purity (%) ^a

Culture broth ^b

Insulin precursor

7041 Da

3075

0.437

100

20

Chelating sepharose FF ^c

Insulin precursor

7041 Da

2921

0.415

95

96

Desalting & Lyophilization

Insulin precursor

7041 Da

2859

0.406

93

98

Transpeptidation

Pre-insulin

5920 Da

n.d. ^d

n.d. ^d

n.d. ^d

n.d. ^d

Resource RPC Desalting & Lyophilization

Pre-insulin

5920 Da

1915

0.323

74

90

Deprotection

Human insulin

5808 Da

n.d. ^d

n.d. ^d

n.d. ^d

n.d. ^d

Resource RPC (two passages), Desalting & Lyophilization

Human insulin

5808 Da

1537

0.264

60

99

^a Recovery and purity was determined by RP-HPLC

^b 200 mg of IP were used for further processing. Data extrapolated to 1-L culture broth

^c Centrifugation step included before chromatography

^d n.d., not determined

Gurramkonda et al. Microbial Cell Factories 2010 9:31   doi:10.1186/1475-2859-9-31