便携式牛乳蛋白质含量检测仪设计与验证
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摘要
为了给牛乳蛋白质含量检测提供一种操作简便、成本低廉的便携式仪器,基于595 nm下考马斯亮蓝G-250染料中的疏水基团在酸性条件下与蛋白质的疏水微区具有较强亲和力,从而影响光透射特性的原理,设计了一种便携式牛乳蛋白质含量检测仪。该检测仪的硬件主要由单片机、光源模块、光传感器模块、输入输出模块和电源模块等组成;软件由Keil C51编写,并由主函数、光照度采集子函数、数据预处理子函数、显示子函数和蛋白质含量计算子函数等组成。以生鲜牛乳为对象,研究了透射光照度与蛋白质含量之间的关系,发现随着蛋白质含量的增加,透射光照度减小,二者具有良好的线性关系,其线性决定系数为0. 872。对比该仪器的蛋白质检测结果表明,同凯氏定氮法相比,本检测仪的绝对误差范围是±0. 08 g/(100 g),平均绝对误差为0. 05 g/(100 g)。此外,该检测仪的检测时间小于2 s,可快速、有效地检测牛乳中蛋白质含量。
Protein content is one of the most important quality indicators of milk. The traditional methods used to measure protein content in milk have high precision,but the measurement procedures are tedious and time-consuming,and the measurement equipments are expensive. In addition,these methods can not be used in field or in-situ measurement. Therefore,developing a kind of portable milk protein content detector with simple operation and low cost,which could be used in-situ measurement,would be appreciated by milk producers,processers and consumers. Since the hydrophobic grouping of Coomassie brilliant blue G-250 has strong affinity for protein in acids,a portable milk protein content detector was developed based on light transmission. The hardware of the detector consisted of the modules of single chip micro-computer,light source,light sensor,input/ouput and power. The software was developed by C51 language,and was made up of main function and several sub-functions to realize light collection,data preprocessing,protein content calculation and display. Raw fresh milk was used to study the relationship between intensity of transmitted light and protein content. It was found that the intensity of transmitted light was decreased with the increase of protein content, and they had strong linear relationship with the coefficient of determination of 0. 872. The validation experiment on the developed detector showed that contrasted with Kjeldahl method,the absolute measurement error of the developed detector was within the range of ± 0. 08 g/( 100 g),and its mean absolute error was 0. 05 g/( 100 g). In addition,the measurement result could be given in 2 s.
Protein content is one of the most important quality indicators of milk. The traditional methods used to measure protein content in milk have high precision,but the measurement procedures are tedious and time-consuming,and the measurement equipments are expensive. In addition,these methods can not be used in field or in-situ measurement. Therefore,developing a kind of portable milk protein content detector with simple operation and low cost,which could be used in-situ measurement,would be appreciated by milk producers,processers and consumers. Since the hydrophobic grouping of Coomassie brilliant blue G-250 has strong affinity for protein in acids,a portable milk protein content detector was developed based on light transmission. The hardware of the detector consisted of the modules of single chip micro-computer,light source,light sensor,input/ouput and power. The software was developed by C51 language,and was made up of main function and several sub-functions to realize light collection,data preprocessing,protein content calculation and display. Raw fresh milk was used to study the relationship between intensity of transmitted light and protein content. It was found that the intensity of transmitted light was decreased with the increase of protein content, and they had strong linear relationship with the coefficient of determination of 0. 872. The validation experiment on the developed detector showed that contrasted with Kjeldahl method,the absolute measurement error of the developed detector was within the range of ± 0. 08 g/( 100 g),and its mean absolute error was 0. 05 g/( 100 g). In addition,the measurement result could be given in 2 s.
引文
[1]张熙桐,关博元,孔繁华,等.人乳、牛乳、羊乳中乳清部分氨基酸组成及乳清蛋白中蛋白质二级结构的对比[J].食品科学,2017,38(24):107-112.ZHANG Xitong,GUAN Boyuan,KONG Fanhua,et al. Comparative study of amino acid composition and secondary structure of whey proteins in human milk,cow milk,and goat milk[J]. Food Science,2017,38(24):107-112.(in Chinese)
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[3] GB 5009. 5—2016食品中蛋白质的测定[S]. 2016.
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[8] NIERO G,PENASA M,GOTTARDO P,et al. Short communication:selecting the most informative mid-infrared spectra wavenumbers to improve the accuracy of prediction models for detailed milk protein content[J]. Journal of Dairy Science,2016,99(3):1853-1858.
[9] MAZUREK S,SZOSTAK R,CZAJA T,et al. Analysis of milk by FT-Raman spectroscopy[J]. Talanta,2015,138:285-289.
[10] HUANG Y,GU J,XIANG G,et al. Detection of total protein in milk using phosphomolybdic acid-mediated surface-enhanced Raman spectroscopy[J]. Journal of Raman Spectroscopy,2016,47(3):277-282.
[11] XIN Q,LING H Z,LONG T J,et al. The rapid determination of fat and protein content in fresh raw milk using the laser light scattering technology[J]. Optics&Lasers in Engineering,2006,44(8):858-869.
[12] RUKKE E O,OLSEN E F,DEVOLD T,et al. Technical note:comparing calibration methods for determination of protein in goat milk by ultraviolet spectroscopy[J]. Journal of Dairy Science,2010,93(7):2922-2925.
[13]孙选.牛奶质量超声检测基础理论及实验研究[D].天津:天津大学,2007.SUN Xuan. Study on theories and experiments of milk quality ultrasonic measurement[D]. Tianjin:Tianjin University,2007.(in Chinese)
[14] HAKODA A,LI Y,NAITO S,et al. Determination of crude protein in macaroni products by the combustion method and comparison with the Kjeldahl method:interlaboratory study[J]. Food Science&Technology Research,2011,17(3):227-232.
[15] AMAMCHARLA J K,METZGER L E. Evaluation of a rapid protein analyzer for determination of protein in milk and cream[J]. Journal of Dairy Science,2010,93(8):3846-3857.
[16]郭文川,康飞,朱新华.频率、温度和大豆蛋白对牛乳介电特性的影响[J/OL].农业机械学报,2015,46(10):274-278.GUO Wenchuan,KANG Fei,ZHU Xinhua. Influence of frequency,temperature and soy protein on dielectric properties of raw milk[J/OL]. Transactions of the Chinese Society for Agricultural Machinery,2015,46(10):274-278. http:∥www. jcsam. org/jcsam/ch/reader/view_abstract. aspx? flag=1&file_no=20151036&journal_id=jcsam. DOI:10. 6041/j. issn.1000-1298. 2015. 10. 036.(in Chinese)
[17] ZHU X,GUO W,KANG F,et al. Determination of protein content of raw fresh cow's milk using dielectric spectroscopy combined with chemometric methods[J]. Food&Bioprocess Technology,2016,9(12):1-11.
[18]郭文川,孔繁荣.不同类型蛋白质及其添加量对牛乳介电特性的影响[J/OL].农业机械学报,2016,47(8):248-254.GUO Wenchuan,KONG Fanrong. Influence of different proteins and their additive amounts on permittivities of milk[J/OL].Transactions of the Chinese Society for Agricultural Machinery,2016,47(8):248-254. http:∥www. j-csam. org/jcsam/ch/reader/view_abstract. aspx? flag=1&file_no=20160832&journal_id=jcsam. DOI:10. 6041/j. issn. 1000-1298. 2016. 08.032.(in Chinese)
[19]丁英强.超声波牛奶成分快速检测方法及仪器研究[D].天津:天津大学,2006.DING Yingqiang. Study on the method and instrument of milk components rapid analysis by ultrasonic[D]. Tianjin:Tianjin University,2006.(in Chinese)
[20] LIN Y,O'MAHONY J A,KELLY A L,et al. Seasonal variation in the composition and processing characteristics of herd milk with varying proportions of milk from spring-calving and autumn-calving cows[J]. Journal of Dairy Research,2017,84(4):444-452.
[21] MUSAAD A M,FAYE B,ALMUTAIRI S E. Seasonal and physiological variation of gross composition of camel milk in Saudi Arabia[J]. Emirates Journal of Food&Agriculture,2011,25(8):618-624.
[22]王永刚,杨光瑞,马雪青,等.荧光光谱法和分子模拟技术研究考马斯亮蓝G-250与牛血清白蛋白的相互作用[J].光谱学与光谱分析,2017,37(8):2474-2479.WANG Yonggang,YANG Guangrui,MA Xueqing,et al. Study on interaction between Coomassie brilliant blue G-250 and bovine serum albumin by fluorescence spectroscopy and molecular modeling[J]. Spectroscopy and Spectral Analysis,2017,37(8):2474-2479.(in Chinese)
[23]乐薇,刘乐.数码成像比色法测定牛乳中蛋白质的含量[J].食品科学,2016,37(4):154-157.LE Wei,LIU Le. Determination of the protein content of milk by digital imaging colorimetry[J]. Food Science,2016,37(4):154-157.(in Chinese)
[24]冯昕,王吉中,尧俊英,等.考马斯亮蓝法测定乳与乳制品中蛋白质含量[J].粮食与食品工业,2010,17(3):57-59.FENG Xin,WANG Jizhong,YAO Junying,et al. Determination of protein content in milk and dairy products by Coomassie brilliant blue method[J]. Cereal&Food Industry,2010,17(3):57-59.(in Chinese)
[25]孙红,陈香,孙梓淳,等.基于透射光谱的玉米叶片含水率快速检测仪研究[J/OL].农业机械学报,2018,49(3):173-178.SUN Hong,CHEN Xiang,SUN Zichun,et al. Rapid detection of moisture content in maize leaves based on transmission spectrum[J/OL]. Transactions of the Chinese Society for Agricultural Machinery,2018,49(3):173-178. http:∥www. jcsam. org/jcsam/ch/reader/view_abstract. aspx? file_no=20180321&flag=1&journal_id=jcsam. DOI:10. 6041/j. issn.1000-1298. 2018. 03. 021.(in Chinese)
[2] GB 19301—2010食品安全国家标准生乳[S]. 2010.
[3] GB 5009. 5—2016食品中蛋白质的测定[S]. 2016.
[4] FENG X D,SU R,XU N,et al. Portable analyzer for rapid analysis of total protein,fat and lactose contents in raw milk measured by non-dispersive short-wave near-infrared spectrometry[J]. Chemical Research in Chinese Universities,2013,29(1):15-19.
[5]王加华,张晓伟,王军,等.基于便携式近红外技术的生鲜乳品质现场评价[J].光谱学与光谱分析,2014,34(10):2679-2684.WANG Jiahua,ZHANG Xiaowei,WANG Jun,et al. On-site evaluation of raw milk qualities by portable vis/NIR transmittance technique[J]. Spectroscopy and Spectral Analysis,2014,34(10):2679-2684.(in Chinese)
[6] BOGOMOLOV A,BELIKOVA V,GALYANIN V,et al. Reference-free spectroscopic determination of fat and protein in milk in the visible and near infrared region below 1 000 nm using spatially resolved diffuse reflectance fiber probe[J]. Talanta,2017,167:563-572.
[7] BONFATTI V,DI M G,CARNIER P. Effectiveness of mid-infrared spectroscopy for the prediction of detailed protein composition and contents of protein genetic variants of individual milk of Simmental cows[J]. Journal of Dairy Science,2011,94(12):5776-5785.
[8] NIERO G,PENASA M,GOTTARDO P,et al. Short communication:selecting the most informative mid-infrared spectra wavenumbers to improve the accuracy of prediction models for detailed milk protein content[J]. Journal of Dairy Science,2016,99(3):1853-1858.
[9] MAZUREK S,SZOSTAK R,CZAJA T,et al. Analysis of milk by FT-Raman spectroscopy[J]. Talanta,2015,138:285-289.
[10] HUANG Y,GU J,XIANG G,et al. Detection of total protein in milk using phosphomolybdic acid-mediated surface-enhanced Raman spectroscopy[J]. Journal of Raman Spectroscopy,2016,47(3):277-282.
[11] XIN Q,LING H Z,LONG T J,et al. The rapid determination of fat and protein content in fresh raw milk using the laser light scattering technology[J]. Optics&Lasers in Engineering,2006,44(8):858-869.
[12] RUKKE E O,OLSEN E F,DEVOLD T,et al. Technical note:comparing calibration methods for determination of protein in goat milk by ultraviolet spectroscopy[J]. Journal of Dairy Science,2010,93(7):2922-2925.
[13]孙选.牛奶质量超声检测基础理论及实验研究[D].天津:天津大学,2007.SUN Xuan. Study on theories and experiments of milk quality ultrasonic measurement[D]. Tianjin:Tianjin University,2007.(in Chinese)
[14] HAKODA A,LI Y,NAITO S,et al. Determination of crude protein in macaroni products by the combustion method and comparison with the Kjeldahl method:interlaboratory study[J]. Food Science&Technology Research,2011,17(3):227-232.
[15] AMAMCHARLA J K,METZGER L E. Evaluation of a rapid protein analyzer for determination of protein in milk and cream[J]. Journal of Dairy Science,2010,93(8):3846-3857.
[16]郭文川,康飞,朱新华.频率、温度和大豆蛋白对牛乳介电特性的影响[J/OL].农业机械学报,2015,46(10):274-278.GUO Wenchuan,KANG Fei,ZHU Xinhua. Influence of frequency,temperature and soy protein on dielectric properties of raw milk[J/OL]. Transactions of the Chinese Society for Agricultural Machinery,2015,46(10):274-278. http:∥www. jcsam. org/jcsam/ch/reader/view_abstract. aspx? flag=1&file_no=20151036&journal_id=jcsam. DOI:10. 6041/j. issn.1000-1298. 2015. 10. 036.(in Chinese)
[17] ZHU X,GUO W,KANG F,et al. Determination of protein content of raw fresh cow's milk using dielectric spectroscopy combined with chemometric methods[J]. Food&Bioprocess Technology,2016,9(12):1-11.
[18]郭文川,孔繁荣.不同类型蛋白质及其添加量对牛乳介电特性的影响[J/OL].农业机械学报,2016,47(8):248-254.GUO Wenchuan,KONG Fanrong. Influence of different proteins and their additive amounts on permittivities of milk[J/OL].Transactions of the Chinese Society for Agricultural Machinery,2016,47(8):248-254. http:∥www. j-csam. org/jcsam/ch/reader/view_abstract. aspx? flag=1&file_no=20160832&journal_id=jcsam. DOI:10. 6041/j. issn. 1000-1298. 2016. 08.032.(in Chinese)
[19]丁英强.超声波牛奶成分快速检测方法及仪器研究[D].天津:天津大学,2006.DING Yingqiang. Study on the method and instrument of milk components rapid analysis by ultrasonic[D]. Tianjin:Tianjin University,2006.(in Chinese)
[20] LIN Y,O'MAHONY J A,KELLY A L,et al. Seasonal variation in the composition and processing characteristics of herd milk with varying proportions of milk from spring-calving and autumn-calving cows[J]. Journal of Dairy Research,2017,84(4):444-452.
[21] MUSAAD A M,FAYE B,ALMUTAIRI S E. Seasonal and physiological variation of gross composition of camel milk in Saudi Arabia[J]. Emirates Journal of Food&Agriculture,2011,25(8):618-624.
[22]王永刚,杨光瑞,马雪青,等.荧光光谱法和分子模拟技术研究考马斯亮蓝G-250与牛血清白蛋白的相互作用[J].光谱学与光谱分析,2017,37(8):2474-2479.WANG Yonggang,YANG Guangrui,MA Xueqing,et al. Study on interaction between Coomassie brilliant blue G-250 and bovine serum albumin by fluorescence spectroscopy and molecular modeling[J]. Spectroscopy and Spectral Analysis,2017,37(8):2474-2479.(in Chinese)
[23]乐薇,刘乐.数码成像比色法测定牛乳中蛋白质的含量[J].食品科学,2016,37(4):154-157.LE Wei,LIU Le. Determination of the protein content of milk by digital imaging colorimetry[J]. Food Science,2016,37(4):154-157.(in Chinese)
[24]冯昕,王吉中,尧俊英,等.考马斯亮蓝法测定乳与乳制品中蛋白质含量[J].粮食与食品工业,2010,17(3):57-59.FENG Xin,WANG Jizhong,YAO Junying,et al. Determination of protein content in milk and dairy products by Coomassie brilliant blue method[J]. Cereal&Food Industry,2010,17(3):57-59.(in Chinese)
[25]孙红,陈香,孙梓淳,等.基于透射光谱的玉米叶片含水率快速检测仪研究[J/OL].农业机械学报,2018,49(3):173-178.SUN Hong,CHEN Xiang,SUN Zichun,et al. Rapid detection of moisture content in maize leaves based on transmission spectrum[J/OL]. Transactions of the Chinese Society for Agricultural Machinery,2018,49(3):173-178. http:∥www. jcsam. org/jcsam/ch/reader/view_abstract. aspx? file_no=20180321&flag=1&journal_id=jcsam. DOI:10. 6041/j. issn.1000-1298. 2018. 03. 021.(in Chinese)