不同物理和化学因素对重组人角质细胞生长因子2稳定性的影响

目的:研究重组人角质细胞生长因子2 (rhKGF-2)在不同保存条件下的稳定性，阐明温度、pH值、缓冲溶液及反复冻融因素对rhKGF-2稳定性的影响作用。方法: rhKGF-2在不同温度(－20℃、4℃、25℃、40℃)、pH值(pH 4～9)、缓冲溶液（柠檬酸钠-柠檬酸缓冲液、磷酸氢二钠-柠檬酸缓冲液、磷酸盐缓冲液、吉斐氏缓冲溶液）及反复冻融条件下分别保存，采用RP-HPLC法检测rhKGF-2纯度，同时观察外观性状。结果：－20℃、4℃分别保存时，rhKGF-2的各项性质在观察期内都无明显变化；25℃时，rhKGF-2存放28 d纯度较0 h(100.00%)下降至(68.20±0.14)%；40℃时，rhKGF-2存放9 h纯度较0 h(100.00%)下降至(4.80±0.39)%。rhKGF-2(pH　6.0)在25℃条件下保存90 d纯度较0 h(100.00%)下降至(79.20±0.12)%，与对照组(0 h)比较差异有显著性(P<0.05)；pH值为4.0、9.0的rhKGF-2液体在25℃条件下分别保存120　h纯度较0 h(100.00%)下降至(15.60±0.30)%和(67.30±0.10)%,与对照组(0 h)比较差异有显著性(P<0.01)。rhKGF-2在柠檬酸缓冲液中25℃条件下保存60　d纯度较0 h(100.00%)下降至(79.20±0.15)%，与对照组(0 h)比较差异有显著性(P<0.01)。rhKGF-2原液反复冻融后其纯度变化差异无显著性(P>0.05)。结论：rhKGF-2在低温和pH 6.0～7.0中性条件下稳定。高温、偏酸性和偏碱性条件下均不利于rhKGF-2稳定。柠檬酸缓冲液有利于rhKGF-2稳定。反复冻融对rhKGF-2稳定性影响不明显。

Effects of different physical and chemical factors on stability of recombinant human keratinocyte growth factor-2

Objective To study the stabilities of recombinant human keratinocytegrowth factor-2(rhKGF-2) deposited at different conditions,and elucidate the influences of temperature,pH,buffer solutions and repeated freeze thawing on rhKGF-2 stability.Methods rhKGF-2 was deposited at different conditions such as temperatures (-20℃,4℃,25℃,40℃),pH(pH 4-9),buffer solutions(Citrate buffer solution,Disodium hydrogen phosphate-Citrate buffer,Phosphate buffer,Jifeishi buffer) and repeated freeze thawing,respectively.The purity of rhKGF-2 was determined by RP-HPLC,and the appearance was also observed.Results All properties of rhKGF-2 were not altered during the observation period at -20℃ and 4℃;the purity of rhKGF-2 deposied for 28 d was reduced to (68.20±0.14)% compared with 0 h(100.00%) at 25℃;the purity of rhKGF-2 deposited for 9 h was reduced to (4.80±0.39)% compared with 0 h(100.00%) at 40℃. The purity of rhKGF-2(pH 6.0) deposited for 90 d was reduced to(79.20±0.12)% compared with 0 h(100.00%) at 25℃(P<0.05);the purities of rhKGF-2(pH 4.0,9.0) deposited for 120 h were respectively reduced to (15.60±0.3)% and (67.30±0.1)% compared with 0 h(100.00%)at 25℃(P<0.01).The purity of rhKGF-2(pH6.0) deposited in Citrate buffer solution deposited for 60 d was reduced to (79.20±0.15)% compared with 0 h(100.00%) at 25℃(P<0.01).The purity of rhKGF-2 had no significant change after repeated freeze thawing(P>0.05).Conclusion rhKGF-2 is stable when deposited at low temperature and pH6.0-7.0 neutral conditions.High temperature,acidic and basic conditions are harmful for rhKGF-2 stability.It is profit for rhKGF-2 stability in Citrate buffer solution.There is no impact on rhKGF-2 stability after repeated freeze thawing.