泌乳期间母乳脂肪球及脂肪酸的变化

蒋文笛，韦伟，王兴国，金青哲**

（江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122）

2.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.02.0State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122；State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122；State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122；State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122;江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122;江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122;江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122214122;214122;;18801512215;15006180870;;18801512215;85329050;;江苏省无锡市滨湖区蠡湖大道1800号;江苏省无锡市滨湖区蠡湖大道1800号;;jiangwendiwendy@126.com;weiw@jiangnan.edu.cn;wxg1002@qq.com;jqzwuxi@163.com蒋文笛(1994),女，硕士研究生，主要研究母乳脂质与营养;韦伟（1988），副教授，新型功能性油脂;;蒋文笛;韦伟;王兴国;金青哲JIANG Wendi;WEI Wei;WANG Xingguo;JIN Qingzhe韦伟 2.02.02.02.02.02.02.02.02.02.02.02.02.02.02.01*|*期刊*|*LANIGAN J. 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Biochimica et Biophysica Acta (BBA) - Reviews on Biomembranes, 1975, 415(3): 273-309.泌乳期间母乳脂肪球及脂肪酸的变化|Changes in fat globules and fatty acids of human milk during lactation||1|蒋文笛|JIANG Wendi|江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122|State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122|蒋文笛(1994),女，硕士研究生，主要研究母乳脂质与营养|江苏省无锡市滨湖区蠡湖大道1800号|214122|jiangwendiwendy@126.com|18801512215|18801512215<CR>*|2|韦伟|WEI Wei|江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122|State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122|韦伟（1988），副教授，新型功能性油脂|江苏省无锡市滨湖区蠡湖大道1800号|214122|weiw@jiangnan.edu.cn|85329050|15006180870<CR>|3|王兴国|WANG Xingguo|江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122|State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122||||wxg1002@qq.com||<CR>|4|金青哲|JIN Qingzhe|江南大学 食品学院，食品科学与技术国家重点实验室，食品安全与营养协同创新中心，江苏 无锡214122|State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122||||jqzwuxi@163.com||

摘要：为探究泌乳初期（30日以内）母乳脂肪球及其脂肪酸组成的变化，本研究采集了无锡地区六位健康产妇泌乳初期（3-30日）的134个母乳样本，分析了样本的粒径、表观电位、脂肪含量、脂肪酸组成及对微观结构进行了观测。研究发现，泌乳期间母乳脂肪球的粒径从3.77±0.95 μm增加到了5.09±0.88 μm。在泌乳初期的几日内（3-12日），脂肪球粒径及其脂肪含量有明显的上升趋势，这可能是因为脂肪的含量与组成发生了变化。初乳样本的中链脂肪酸、超长链脂肪酸及饱和脂肪酸成分含量较多，而长链脂肪酸与不饱和脂肪酸含量较少。因此，初乳含较低含量的乳脂，脂肪球粒径较小，具有高含量的中链脂肪酸及多不饱和脂肪酸，对婴儿的生长发育可能有特殊的益处

关键词：人乳脂; 人乳脂肪球; 脂肪酸组成; 泌乳阶段; 粒径分布

中图分类号：TS252

Changes in fat globules and fatty acids of human milk during lactation

JIANG Wendi, WEI Wei, WANG Xingguo, JIN Qingzhe

(State Key Laboratory of Food Science and Technology, Collaborative innovation center of food safety and quality control in Jiangsu Province, School of Food Science and Technology, Jiangnan University, Wuxi 214122)

Abstract: We aimed to gain new insights into the variations of human milk fat globules (HMFGs) and the association with the fatty acid (FA) composition during lactation. HM samples from six healthy Chinese mothers were collected every day from 3 days to 30 days postpartum (n = 134). The size distribution of HMFGs in HM samples were investigated using a laser particle size analyzer and confocal laser scanning microscopy (CLSM). FA composition of triacylglycerol (TAG) was also analyzed. Size distribution and CLSM images indicated that the size of HMFGs increased during lactation from 3.77±0.95 μm to 5.09±0.88 μm. A significant increase was observed in the size of HMFGs and fat content in HM samples for the first 10 days of lactation (colostrum). Changes in the size of HMFGs during lactation may have been associated with the content and composition of fat. Colostrum samples showed higher content of middle-chain fatty acids (MCFAs), very-long-chain fatty acids and saturated fatty acids, but lower content of long-chain fatty acids and unsaturated fatty acids. These results suggest that colostrum had low fat content, relatively small HMFGs, and high content of MCFAs and polyunsaturated fatty acids, which may confer special benefits for the needs of newborns.

Key words: Human milk fat, human milk fat globules, fatty acid composition, lactation stage, size distribution

引言

母乳被广泛认为是婴儿最为理想的食品[1]，尤其是对于6个月以内的新生儿。母乳因含有营养物质与其他非营养的微量物质，影响着婴儿的健康和生长[2]。母乳通常可分为初乳（产后1-6天），过渡乳（产后7-14天）和成熟乳（产后15天以后），其中脂肪是母乳最有价

值的主要成分[3]。

母乳中脂肪的比例约为3 ％-5 ％，提供母乳喂养婴儿所需能量的50 %。母乳脂肪以复合三层膜包裹内核的脂肪球结构存在于水包油形式的乳液中，称为人乳脂肪球（HMFG）。人乳脂肪球的外部三层膜结构富含磷脂及蛋白质，而内核为三酰基甘油酯[4]，具有广泛的粒径分布（0.2-15 μm，平均粒径3-5 μm）。

近年来，关于泌乳期间人乳脂肪变化的研究逐渐增多。在泌乳期间，人乳脂肪发生了显著的变化，包括脂肪含量[5]，脂肪酸组成[6,7]，三酰基甘油酯的组成[8,9]和生育酚[10]的变化。一些研究已经报道了泌乳期间母乳与牛乳脂肪球粒径以及脂肪酸组成的差异[11-17]，但研究结论并不一致。据报道，随着泌乳期的发展，乳脂肪球的平均粒径体现出了没有显着性差异[12,18]，减少[13,15,16]和增加[11,14,17]的规律，这可能是由于样品的储存或处理以及分析方法的差异[15]。此外，泌乳期间人乳脂肪球结构的变化尚不清楚，这对于人乳脂肪和新型婴儿配方奶粉的开发极为重要。

本研究收集了6位产妇产后第3天至第30天的134份母乳样本，研究了不同泌乳时期人乳脂肪球的粒径分布、脂肪含量与zeta表观电位，还研究了泌乳期间与人乳脂肪球粒径相关的脂肪酸组成的变化。本研究获得的数据有助于婴儿配方奶粉的研究和开发。当母乳喂养不充分或无条件进行母乳喂养时，将需要在化学组成和物理结构方面与母乳更为相似的婴儿配方乳粉作为婴儿的食物供给。

材料与方法

样本与试剂

本研究的134个母乳样本由无锡地区六位健康的产妇捐赠，分为初乳（3-6天），过渡乳（7-14天）和成熟乳（15-30天）。样本于两小时内进行分析检测，提取脂肪并存于-80 ºC进行后续脂肪酸组成分析。6位产妇的主要信息如下（表1）。

表1 产妇及新生儿的基本信息

Tab. 1 Basic information of mothers and infants

各项指标	数值
生产方式（顺产/剖腹产）	6 / 0
年龄（岁）	28.00 ± 3.56
身高（cm）	162.00 ± 3.96
体重（kg）	58.50 ± 7.39
孕期增重（kg）	16.58 ± 6.09
胎儿体重（kg）	3.35 ± 0.21
胎儿性别（女/男）	4 / 2

注：数据表达为平均值±标准方差

尼罗红荧光染料（9-diethylamino-5H-benzoalha-phenoxazine-5-one ，0.1 mg/mL溶于丙酮; Sigma-Aldrich，美国），荧光头基标记的磷脂N-（丽丝胺罗丹明B磺酰基）二油酰磷脂酰乙醇胺（Rh-DOPE，1 mg / mL溶于氯仿；Avanti Polar Lipids，美国）。37种脂肪酸甲酯（FAME Mix，C4-C24）的标准品和正己烷（> 99.9 ％纯度）分别从Sigma-Aldrich和J＆K Scientific（中国北京）获得。所有其他试剂和溶剂均为分析级，购自国药化学试剂（中国上海）。

实验方法

粒径分布

S3500型激光粒度分析仪（美国Microtrac公司）测定母乳脂肪球粒径分布。选用湿法模式测定，样品分析参数设定为球形，折射系数1.460。在0.2 mL样本中依次加入0.2 mL EDTA/NaOH（35 mM，pH=7.0）缓冲液与0.2 mL SDS溶液（1 %）来裂解酪蛋白胶束与防止脂肪球之间的聚集。上述溶液混合后进行测定，每个样品重复3次以上取平均值。

zeta表观电位

利用纳米粒度及zeta电位分析仪（英国Malvern公司）在25 ºC下测定样品的zeta表观电位。在3-5 μL样品中加入10 mL缓冲液 （5 mM CaCl2, 50 mM NaCl, 20 mM 咪唑，pH=7.0），每个样品重复3次以上取平均值。

脂质含量

脂质的提取参考了Röse–Gottlieb[3]的方法。5 mL样品中加入1 mL氨水，充分混匀，并于65 ± 5 ºC震荡水浴，15-20 min后取出冷却至室温。依次加入5 mL无水乙醇、12.5 mL无水乙醚与12.5 mL石油醚，充分震荡混匀。30 min后待混合物静置分层，分离上清液。上述操作重复两次，合并上清液，氮气吹干至恒重，称量所得脂质。脂质含量按下式计算所得：

脂质含量（%）=乳脂质量（g）/母乳样品质量（g）×100

脂肪酸组成

取10 mg脂质，加入700 μL正己烷与125 μL KOH-CH3OH（2 mol/L），旋涡振荡混匀后，加入适量无水硫酸钠混匀，静置，取上清过膜后进行分析。利用配备有Trace TR-FAME毛细管柱（60 m×0.25 mm×0.25 μm）、氢火焰离子检测器与7820A系统的气相色谱（GC）进行脂肪酸组成分析。检测条件：检测口与进样口温度设定为250 ºC；载气流速为1.2 mL / min，分流比为1:20；柱箱温度在60 ºC维持3分钟后，以5 ºC/分钟的速率升至175 ºC，保持15分钟，然后以2 ºC/分钟的速率升至220 ºC并保持10分钟。比较样品中包含的脂肪酸甲酯（FAME）的峰与37种FAME标准品出峰的保留时间来确定脂肪酸组成。

激光共聚焦显微镜

选用LSM-710型激光共聚焦显微镜（德国Zeiss公司）进行样品的微观结构观测。0.1 mg/mL Nile Red染料与1 mg/mL Rh-DOPE染料分别与样本以1:10与1:40比例混合，避光染色30 min以上，取8 μL于载玻片上，20 μL的琼脂糖（0.5 % w/v，45 °C）固定，盖上盖玻片。Nile Red于488 nm 处，Ar+激光器激发，发射光接收范围500-600 nm，Rh-DOPE于552 nm处，He-Ne激光器激发，发射光接收范围570-670 nm，利用×40物镜进行观测。

统计分析

本文数值表示为平均值±标准偏差，除粒径分布数据重复5次测定以外，其余实验重复测定三次。采用IBM SPSS软件（20.0版，IBM，Armonk，NY，USA），ANOVA的Ducan检验法进行单因素方差分析（α > 0.05判定为方差齐性，p < 0.05时具有差异，p<0.01时差异显著）。

结果与讨论

人乳脂肪含量

采集母乳的脂肪含量处于3-5 %之间。从第3日至第11日，样本脂质含量由3.24 ± 1.68 %增加到了4.96 ± 2.13 %，之后则保持平稳（图1a）。如图1b中箱型图所示，成熟乳的脂质含量（3.89 ± 1.52 %）处于初乳（3.07 ± 1.03 %）及过渡乳（4.01 ± 1.71 %）的脂质含量之间。自初乳至过渡乳分泌期间，脂质含量增加显著（p < 0.01），但之后的脂质含量未有明显的变化（p = 0.468），这在一些研究中也有相似的报道[19-22]。另外，一些研究表明，在泌乳期间母乳脂质含量持续明显地上升[15,23]，而另一些研究显示，不同阶段母乳之间脂含量差异并不显著[24]。

图1 泌乳期间母乳脂质含量的变化（** p < 0.01, * p < 0.05）

Fig. 1 Changes of human milk fat content as lactation progress

人乳脂肪球的粒径分布

图2a显示了母乳脂肪球的体积平均粒径随泌乳期的变化。第3日（初乳）至第12日（过渡乳）期间，体积平均粒径显著上升，分布范围处于3.77 ± 0.95 μm至5.09 ± 0.88 μm之间，之后稳定在5 μm左右。

图2 泌乳期间人乳脂肪球（a）平均粒径及（b）zeta表观电位值的变化

Fig. 2 Changes of HMFG average diameters (a) and zeta potential value (b) during the day of lactation.

最初12日以内母乳脂肪球粒径分布呈现在图3中。曲线逐渐由双峰的分布模式（初乳）转变为三峰的分布模式（过渡乳），且初乳的脂肪球粒径分布范围（平均粒径3.27-3.89 μm）要比过渡乳的分布范围广（4.62-5.50 μm）。总而言之，在泌乳期间，存在大粒径的脂肪球比例增多且样本体积平均粒径增加的现象。

图3 泌乳初期10日内母乳中脂肪球粒径分布的变化

Fig. 3 Milk fat globule size distribution changes of human milk samples during the first 10 days of lactation.

上述结果与一些学者的研究结果类似[25,26,18]。其中，Harzer，Jensen[25,26]等人提出，母乳脂肪球的粒径在泌乳最初四周内有上升的现象，而Rüegg[18]等发现自初乳分泌至此后4-5个月期间，脂肪球粒径从约1.80 μm持续增加至约4.00 μm。还有研究表明，一位依靠人工挤乳的母亲在分泌成熟乳期间脂肪球粒径也体现出增加趋势[12]。

我们的研究发现，泌乳期间母乳样本中的脂质含量随脂肪球平均粒径增加而增加（图2a,1a），与之相似的结论也曾被报道[25,26]。在牛乳中，乳脂肪球的平均粒径也与日泌乳量存在联系[11]。这是因为，乳脂分泌增加时，泌乳细胞分泌的包裹脂滴的膜材料没有同步增加，因此脂滴在被质膜包裹前积聚形成了较大的脂滴。Whittlesto等人[27]并没有观察到母乳与牛乳中脂肪球平均粒径与乳脂含量之间存在的联系，这可能是不同的分析手段与样本所导致的。

人乳脂肪球的zeta表观电位

乳脂肪球表面的电位不能被直接测量，但zeta表观电位（即距微粒表面一定距离的表面电位）可被测定[28]。图2b中显示，初乳与过渡乳之间zeta表观电位的差异较显著（p < 0.01），整个泌乳阶段zeta表观电位的范围处于-6 mV 至 -7.5 mV之间。据Michalski等人[29]报道，母乳成熟乳的脂肪球zeta表观电位在-7.8 ± 0.1 mV左右，比牛乳中脂肪球的表观电位值要低（-13.5 ± 0.9 mV）。另有文献报道母乳脂肪球zeta表观电位平均在-7.6 ± 0.5 mV[30]。我们的研究数据表明，泌乳阶段的最初11日之内，电位值存在下降的趋势（范围处于-6.23 ± 1.14 mV与-7.32 ± 0.63 mV间），而之后的阶段中保持平稳。有文献表明[15]，不同泌乳阶段间人乳脂肪球zeta表观电位的变化是由于水相环境中不同的蛋白质、极性脂与矿物质所导致的。具有高zeta表观电位的脂肪球更稳定，不易凝集，而表观电位较低的脂肪球对于新生儿来说更利于脂质消化。

人乳脂肪酸组成

不同泌乳时期母乳样本的脂肪酸组成是不同的，母乳脂质的特性主要取决于其脂肪酸组成。有文献表明[25,31]，每日晨间与晚间的母乳样本无显著性差异。下表2为初乳、过渡乳与成熟乳样本的脂肪酸组成。

表2 初乳、过渡乳与成熟乳样本的脂肪酸组成（wt %）

Table 2 Fatty acid composition of colostrum, transitional and mature milk (wt %)

Fatty acid	Colostrum	Transitional milk	Mature milk
(n=21)	(n=43)	(n=70)
10:0	1.18 ± 0.77	1.32 ± 0.59	0.91 ± 0.40b,c
12:0	5.35 ± 2.13	4.99 ± 2.00	3.27 ± 1.28b,c
14:0	5.67 ± 1.23	4.46 ± 1.70a	3.18 ± 1.21b,c
15:0	0.16 ± 0.02	0.14 ± 0.05	0.14 ± 0.06
16:0	21.63 ± 1.49	20.47 ± 1.64a	20.73 ± 2.10
17:0	0.29 ± 0.04	0.28 ± 0.04	0.27 ± 0.05
18:0	5.08 ± 0.68	5.37 ± 0.88	5.88 ± 0.95b,c
20:0	0.76 ± 0.22	0.52 ± 0.08a	0.47 ± 0.07b
22:0	0.71 ± 0.35	0.54 ± 0.20a	0.44 ± 0.12b,c
16:1 n-7	2.49 ± 0.48	2.52 ± 0.53	2.52 ± 0.47
17:1 n-7	0.15 ± 0.03	0.15 ± 0.03	0.17 ± 0.08
18:1 n-9	30.92 ± 3.04	31.55 ± 2.65	34.32 ± 2.65b,c
18:2 n-6	22.28 ± 2.24	24.49 ± 3.45a	24.57 ± 3.36b
18:3 n-3	1.38 ± 0.31	1.63 ± 0.46a	1.55 ± 0.48
18:3 n-6	0.41 ± 0.11	0.35 ± 0.07a	0.38 ± 0.07
20:3 n-6	0.91 ± 0.30	0.76 ± 0.15a	0.74 ± 0.15b
22:6 n-3	0.62 ± 0.47	0.46 ± 0.22a	0.48 ± 0.27b
ΣSFA	40.83 ± 3.61	38.10 ± 4.10a	35.28 ± 3.70b,c
ΣMUFA	33.57 ± 3.37	34.22 ± 2.83	37.01 ± 2.70b,c
ΣPUFA	25.61 ± 2.60	27.70 ± 3.76a	27.71 ± 3.72b
n-6/n-3	11.85 ± 2.45	12.32 ± 2.29	12.83 ± 2.52
LA/ALA	16.11 ± 2.58	15.17 ± 2.37	16.12 ± 3.17

注：a 初乳与过渡乳之间存在显著性差异（p < 0.05），b 初乳与成熟乳之间存在显著

性差异（p < 0.05），c 过渡乳与成熟乳之间存在显著性差异（p < 0.05）；LA/ALA：亚

油酸与亚麻酸相对含量的比例。

母乳样本中检测出了17种脂肪酸，其中包括9种主要脂肪酸（相对含量> 95 %）与8种微量脂肪酸（相对含量< 1 %）。这其中仅有一种奇数碳脂肪酸（17:1 n-7）与两种反式脂肪酸（18:1 n-9与16:1 n-7）被检测出。

样本检测出了9种饱和脂肪酸，其中初乳中占比40.83 ± 3.61 %而成熟乳中占比35.28 ± 3.70 %，在泌乳期间相对含量显著降低（p < 0.01）。图4为饱和脂肪酸（SFAs）、单不饱和脂肪酸（MUFAs）与多不饱和脂肪酸（PUFAs）在泌乳期间样本中相对含量的箱形图。在所有的SFAs中，棕榈酸（16:0）占比约50 %，初乳阶段至过渡乳阶段，含量显著降低（p < 0.05），文献中对于中国与西班牙的母乳样本中曾有相似报道[32-34]。在泌乳阶段，18:0是唯一一种自泌乳初期至成熟乳阶段含量显著上升的脂肪酸（p < 0.05），相似的结论对于我国及其他国家的母乳样本也有报道[31,32,34]。就不同粒径的脂肪球的脂肪酸组成对比来说[35,36]，18:0在粒径较大的脂肪球中含量更多，对于15:0与17:0两种奇数碳脂肪酸，整个泌乳过程的含量较为稳定。一些文献报道[15,31]，随着乳汁的成熟，SFAs的总含量没有明显差异。

图4 不同泌乳时期母乳饱和脂肪酸、单不饱和脂肪酸与多不饱和脂肪酸含量的变化（** p < 0.01）

Fig. 4 Box plots of saturated fatty acid (SFA), monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) in human milk during the lactation (** p < 0.01).

样本的单不饱和脂肪酸（MUFAs）中，我们观察到油酸（18:1 n-9）含量明显上升了（p < 0.05），在成熟乳中含量高达34.32 ± 2.65 %（表2）。这与Chen，Sala-Vila等人[32,34]的结论相似，但与另一些报道的变化趋势略有不同[31,33]。尽管18:1 n-9非必需脂肪酸，但它降低了甘油三酯的熔点，为脂肪球提供了流动性，从而进行组合、运输与新陈代谢等，是结构成分与能量的来源[37]。除了脂肪酸18:1 n-9，我们并没有观察到16:1 n-7 与17:1等明显的变化。若考虑不同粒径脂肪球的脂肪酸组成，有文献报道，油酸（18:1 n-9）在大粒径的脂肪球中含量更高[36]，但另有学者认为，相比于小粒径的脂肪球，大粒径脂肪球中18:1 n-9含量反而降低了[38]，这也可能是脂肪提取方法或是检测到的脂肪酸种类各异所造成的。

相比于过渡乳与成熟乳，初乳中所含多不饱和脂肪酸（PUFA）较少（图4），但由过渡乳向成熟乳转变期间，PUFA含量几乎没有变化（表2）。22:6 n-3（DHA）有助于婴儿智力的提升[15]，并且可能对视网膜功能与髓鞘形成有利。它在初乳中含量相对较高，可能与细胞膜中长链脂肪酸分子含量较为丰富有关，Agostoni[39]，Koletzko[40]，Nyuar[41]及Wang[42]等学者相继有过相似的报道。而Qi等学者[43]还曾报道，无锡地区的母乳样本相比其他一些地区，DHA含量相对更高，这可能是食用水产品的饮食习惯所导致的。

泌乳时期n-6与n-3脂肪酸的比例随时间相对增加。对于必需脂肪酸来说，亚油酸（LA, 18:2 n-6）含量最高，约占总脂肪酸的22.28 ± 2.24 % 至 24.57 ± 3.36 %，另一必需脂肪酸，α-亚麻酸（ALA, 18:3 n-3），在过渡乳与成熟乳中含量相对丰富，而γ-亚麻酸（GLA, 18:3 n-6）与20:3 n-6则在初乳中含量较高，之后出现降低（p < 0.05）。在初乳与过渡乳中，LA:ALA比例有些许降低，但差异并不显著，同时，Lópezlópez等人[33]也发现LA:ALA比例在初乳中偏低。值得提及的是，LA与ALA这两种脂肪酸确保了足够的生物学必需脂肪酸来源，为婴儿提供营养，相关标准中规定[44]，LA:ALA比例应处于5:1至15:1之间。但由于全球其他一些地区母乳中LA:ALA的比例也未达标准[43]，因此，欧盟2016年的最新标准中，删除了这一项。

总的来说，SFAs含量自40.83 ± 3.61 %持续下降到35.28 ± 3.70 %的同时，MUFAs与PUFAs含量分别显著上升（p < 0.01）至37.01 ± 2.70 %（成熟乳）与27.70 ± 3.76 %（过渡乳）。对于不同链长的脂肪酸，长链脂肪酸（LCFAs）含量上升而中链（MCFAs）与超长链脂肪酸（VLCFAs）含量则下降（图5）。图6中的箱形图显示，泌乳的不同阶段之间MCFAs与LCFAs具有显著性差异。与初乳相比，VLCFAs在过渡乳与成熟乳中显著降低了。有文献表明[36,45,46]，小粒径脂肪球中，MCFAs含量较丰富，这也显示了乳腺细胞的生物合成能力的变化[47]。

图5 母乳中链脂肪酸、长链脂肪酸与超长链脂肪酸含量随泌乳期的变化

Fig. 5 The content of middle-chain fatty acid (MCFA), long-chain fatty acid (LCFA) and very long-chain fatty acid (VLCFA) in human milk fat during the day.

图6 不同泌乳时期母乳中链脂肪酸、长链脂肪酸与超长链脂肪酸含量的变化（** p < 0.01, *p < 0.05）

Fig. 6 Box plots of MCFA, LCFA and VLCFA in human milk fat duing the lactation (** p < 0.01, *p < 0.05).

随着泌乳的进程，母乳样本脂肪酸组成的变化，可能是由于新生儿的需求变化所导致的。总的来说，我们的结论反应出母亲的新陈代谢适应性，并且不同粒径大小的脂肪球组成可能不同，类似假设在相关文献中也有提及[36,46,48,49]。

人乳脂肪球的微观结构

泌乳前7日之内的母乳样本经尼罗红与Rh-DOPE荧光染料染色后在激光共聚焦显微镜下观测（初乳与过渡乳样本）。图7中显示了泌乳第3、5、7天的母乳脂肪球。尼罗红染色的中性脂（TAG）呈现于脂肪球的核心，而由Rh-DOPE标记的磷脂则包裹于甘油三酯外侧呈圆环状。这是因为脂肪球分泌时，脂滴由内质网分泌进入细胞质，覆盖了三层膜的包裹，再被分泌出细胞[4]。图中，圆环上的荧光强弱不一，这可能是磷脂分布不均匀所导致的。

图7 激光共聚焦显微镜下泌乳第3（a, a’）、第5（b, b’）、第7（c, c’）天的母乳样本，非极性脂由尼罗红染色（a, b, c），极性脂由Rh-DOPE染色（a’, b’, c’）

Fig. 7 Images of human milk fat globules observed under CLSM for the 3 d (a, a’), 5 d (b, b’) and 7 d (c, c’). Lipids (a, b, c) were labelled by Nile red, and polar lipids (a’, b’, c’) was labelled by Rh-DOPE fluorescent probe.

泌乳最初的7日内，乳脂肪球的粒径显著增加（图7），这与激光粒度仪测定的母乳样本的粒径分布结论一致（图2、3），都显示大粒径脂肪球随泌乳期延长比例增加。在最初的泌乳时期，母乳脂肪球的体积平均粒径增加，这是首次由激光共聚焦显微镜观测到的母乳脂肪球粒径的增加。

Wiking，Rüegg等人[11,18]发现，在牛乳中，乳脂含量，脂肪酸组成与含量，γ-谷氨酰转肽酶的活性，激素的减少（例如，催产素，催乳素）与脂肪球膜均与脂肪球的粒径相关。在人乳中也有相似结论得出[50,40]，但另一方面，Zou等人[15]表示，由于不同的基因或是饮食习惯不同，也可能有不同结论的提出。

结论

本研究收集了无锡地区产妇最初泌乳时期第3至30天的母乳样本，并对其脂肪球粒径、脂肪酸组成变化等指标进行研究。研究数据表明，随泌乳期延长，母乳脂肪球粒径存在增加的趋势。在最初十日内，脂质含量也随着大粒径脂肪球比例的增加而提升，而zeta表观电位却趋向于降低，这些指标在之后维持稳定。初乳相比于过渡乳与成熟乳，小粒径的乳脂肪球占多数，饱和脂肪酸（SFAs），中链脂肪酸（MCFAs）与超长链脂肪酸（VLCFAs）含量丰富，但不饱和脂肪酸（UFAs）与长链脂肪酸（LCFAs）含量较低。本研究以新视角切入研究了母乳脂肪球随泌乳期的影响，未来将有更多的研究着重于揭示其与新生儿机能方面的联系。本文的研究数据有利于婴幼儿配方产品的研发，使其无论是物理结构还是化学组成更贴近于母乳，从而在缺失母乳喂养时为新生儿提供更适合的膳食。

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作者简介：蒋文笛(1994),女，硕士研究生，主要研究母乳脂质与营养

通信联系人：韦伟（1988），副教授，新型功能性油脂. E-mail: weiw@jiangnan.edu.cn