亚硝基血色原的提取与分离方法

The Extraction and Isolation Method of Nitrosyl Hemochromogen

【目的】筛选获取纯度较高的蒸煮腌肉主导色素亚硝基血色原(nitrosohemochromogen,NH)的方法。【方法】采用7种不同极性溶剂对NH进行提取,分析提取液的UV吸收特性和电子顺磁共振光谱(EPR)特性以及残渣的色调。【结果】石油醚完全不溶解NH;丙酮对NH的提取率很高,而且提取液相对稳定;乙酸乙酯提取率不高,但可以保持NH的粉红色。而直接用丙酮提取的NH,EPR光谱检测不到NH的特征信号,检测结果却是脂肪氧化形成自由基的信号。为此,建立了一种三步顺序提取和分离NH的方法,可有效排除脂肪的干扰和氧化,在110 K的温度环境下,EPR光谱可以清晰检测到NH血红素卟啉Fe原子与-14NO基团中的14N原子耦合产生的超精细分裂信号。整个操作在暗室吹氮气的通风橱中进行。将蒸煮腌肉切丁,先用3倍的石油醚(沸程为30—60℃)在黑暗、无氧条件下浸提36 h,弃掉石油醚,然后加入1.5倍的丙酮提取10 min,过滤得到亮红色溶液,蒸发丙酮后再用乙酸乙酯分次萃取,除去乙酸乙酯,即可得到NH。【结论】采用石油醚、丙酮、乙酸乙酯三步顺序提取和分离NH可以防止脂肪氧化对NH的影响,是一种有效的提取和分离NH的方法。

[ Objective ] The purpose of the experiment is to obtain high purity nitrosyl hemochromogen （NH）. [ Method] Seven kinds of different polarity organic solvents were used to extract and isolate the Nit. The UV and EPR spectral characteristics of extracted solution and the hue of meat residue after being extracted NH were analyzed. [ Result ] NH is insoluble in petroleum ether completely. Acetone can dissolve NH efficiently and the extracted solution is relatively stable. Extraction rate of ethyl acetate is not high, but which can keep the pink of NH. The EPR spectrum of NH extracted by acetone was showed the signal information of free radical not of the NH because of the presence of the lipid. For these results, a three-step gradational extraction and isolation procedure with petroleum ether/acetone^ethyl acetate step by step was established based on the characteristics of UV and EPR spectroscopy of NH solution. The diced sample was extracted with 3 times the petroleum ether （ boiling range is 30-60~C） in the dark, anaerobic conditions for 36 h, discard the petroleum ether, and then add 1.5 times the acetone extraction for 10 rain, filter to obtain bright red solution. The acetone was removed and the NH was isolated with ethyl acetate again. [ Conclusion ] It indicated that the three-step gradational extraction procedure with petroleum ether/acetone/ethyl acetate step by step is an effective method for isolation of NH.