GPC-HPLC法测定南极磷虾油中虾青素及校正因子计算

以保健食品南极磷虾油为研究对象,建立了测定南极磷虾油中虾青素含量的全自动凝胶渗透色谱(GPC)净化-高效液相色谱分析方法。样品采用GPC净化,NaOH甲醇溶液皂化后,经C30液相色谱柱分离后测定。考察了皂化溶剂、加碱量、皂化时间等对虾青素皂化效率的影响以及pH对溶液稳定性的影响,并对虾青素异构体的校正因子进行了计算,确定了虾青素的定量方式。南极磷虾油中虾青素的定量限为0.5 mg/kg;在0.1～5 mg/L时峰强度与质量浓度的线性关系良好(r>0.999);加标回收率为96%～98.5%;相对标准偏差为3.0%～5.6%。方法适用于南极磷虾油中虾青素的实际检测。     

NaOH皂化反相高效液相色谱法测定雨生红球藻中虾青素

建立了一种雨生红球藻中虾青素酯皂化的反相高效液相色谱方法。采用二极管阵列检测器（DAD）在476nm处进行检测,虾青素在15min内得到了较好的分离。虾青素的含量在1～15mg／L范围内与峰面积具有良好的线性关系,相关系数为0．9993;方法的检出限为0．049mg／L,RSD为3．27％,及回收率为100．7％。应用本法测定出雨生红球藻中虾青素的总含量为9．79～24．70mg／L。     

高效液相色谱–光电二极管阵列法测定虾青素的含量

建立虾青素含量测定的高效液相色谱–光电二极管阵列法。采用Purospher STAR RP 18(4.6 mm×250mm,5μm)色谱柱,以甲醇–水(体积比为95∶5)为流动相,流速1.0 mL/min,检测波长为482 nm,柱温为30℃,进样量为20μL。在所选定的液相色谱条件下,虾青素主峰与其它杂质峰分离良好,虾青素在0.2~16μg/mL范围内线性良好,线性相关系数r=0.999 9,检出限为0.01μg/mL,测定结果的相对标准偏差为0.42%(n=6),平均回收率为100.4%。该法分析快速准确、灵敏度高、重现性好。     

雨生红球藻中虾青素的C_(30)-反相高效液相色谱法测定

建立了雨生红球藻中虾青素的C30-反相高效液相色谱测定方法。雨生红球藻经二氯甲烷-甲醇(体积比25∶75)研磨和超声提取后,在0.5 mL 0.1 mol/L NaOH甲醇溶液中于5℃避光皂化12 h,在YMC-Ca-rotenoid C30色谱柱(250 mm×4.6 mm×5μm)上以甲醇(A)、叔丁基甲基醚(B)、1%磷酸溶液(C)为流动相梯度洗脱,采用紫外检测器在474 nm测定。在优化实验条件下,雨生红球藻中的虾青素酯可以很好地转化为游离虾青素。实验以虾青素酯和虾青素在C30色谱柱上不同的出峰时间作为判断是否皂化完全的依据。在0.1~5mg.L-1时峰强度与虾青素质量浓度呈良好线性(r=0.999 7),方法的定量下限为2 mg/kg,平均回收率为95%~108%,相对标准偏差为3.5%~14.7%。方法准确可靠,适用于雨生红球藻中虾青素含量的测定。     

超高效液相色谱法测定虾青素

建立超高效液相色谱法快速检测虾青素的方法。采用UPLC BEH C_8色谱柱(50 mm×2.1 mm,1.7μm),考察了流动相、流量及柱温对虾青素样品分离的影响,确定了最佳色谱条件:等度洗脱,流动相为甲醇–水(体积比为75∶25),流量为0.5 mL/min,柱温为40℃,检测波长为475 nm。虾青素的质量浓度在0.2~10.0μg/mL范围内与其色谱峰面积呈良好的线性关系,线性相关系数r=0.998 8,检出限(S/N=3)为0.1μg/mL,定量限(S/N=10)为0.2μg/mL。测定结果的相对标准偏差为0.41%(n=6),加标回收率为105.8%~110.3%。该方法快速、简单、可靠、灵敏、重复性好,可用于虾青素有关样品的快速检测。     

测定雨生红球藻中虾青素及其它色素含量的高效液相色谱法

建立了一种分离和测定雨生红球藻中虾青素和其它色素的反相高效液相色谱法 ;含有雨生红球藻细胞的培养液经离心分离 ,匀浆破碎 ,用甲醇 -二氯甲烷 (体积比3∶1)提取色素 ,提取液用HPLC分离测定 ;分析柱为Nova_PakC18(300mm×3.9mmID ,5μm)柱 ,流动相为水、甲醇和丙酮 ,流速1.0mL/min ,用二极管阵列检测器在250～700nm波长范围扫描 ,在476nm处进行检测 ;虾青素、多种类胡萝卜素和叶绿素在30min内得到较好分离 ,其中游离虾青素、斑蝥黄质和 β_胡萝卜素的检出限分别为3.56、1.36和29.4μg/L,这3种类胡萝卜素分别在质量浓度为1.99～31.7mg/L、2.44～39.0mg/L、2.25～36.0mg/L范围内与峰面积线性关系良好 ,相关系数分别为0.9993、0.9987和0.9778;该法的精密度 (RSD为0.09 %～2.97% )和回收率(96%～102 % )均符合方法学要求 ,可以用于研究和测定雨生红球藻中虾青素和其它色素的含量     

高效液相色谱法测定油脂类保健食品中虾青素

提出了高效液相色谱法测定油脂类保健食品中虾青素含量的方法。样品经乙腈提取,提取液浓缩后,用Dikma C18色谱柱(4.6mm×250mm,5μm)分离,用不同配比的乙腈和水的混合溶液为流动相梯度洗脱,用紫外检测器于波长479nm处检测。虾青素的质量浓度在30.0～300mg.L-1范围内与其峰面积呈线性关系,检出限(3S/N)为0.07mg.L-1。在空白样品中进行加标回收试验,方法的回收率在90.0%～96.7%之间;方法的相对标准偏差(n=6)为2.8%。     

固相萃取-反相高效液相色谱法同时测定饲料中的角黄素和虾青素

建立了一种同时测定饲料中角黄素和虾青素的固相萃取-高效液相色谱法（HPLC）。样品由乙腈提取,经LC-NH2固相萃取小柱净 化,洗脱剂为乙腈-甲苯（体积比为3∶1）,洗脱液被浓缩后进行HPLC分析,色谱柱为ZORBAX Eclipse XDB-C18柱(150 mm×4.6 mm,5 μ m),流动相为乙腈-甲醇（体积比为95∶5）,流速1.0 mL/min,采用二极管阵列检测器检测,检测波长为474 nm;外标法定量。角黄素和 虾青素的线性范围分别为1.0～30.0 mg/L和1.0～20.0 mg/L,相关系数分别为0.9990和0.9991,回收率为90%～101%,相对标准偏差为 0.62%～3.68%,检出限分别为0.84和0.60 mg/L。该方法简便、快速、准确,可用于饲料中角黄素和虾青素的同时测定。     

超高效液相色谱串联质谱法快速检测烤鳗虾中硝基呋喃代谢物残留新技术

采用高检测灵敏度超高效液相色谱串联质谱开发了烤鳗虾中硝基呋喃代谢物残留量快速检测新技术.该技术包括2-氯苯甲醛为衍生剂,试剂盒方法快速样品制备,高检测灵敏度超高效液相色谱串联质谱快速测定.设计了4个添加水平(0.2、0.5、1.0、2.0 μg/kg)、8次重复的试验.结果表明,方法线性范围0～5.0 μg/kg;检测限均为0.2μg/kg;AOZ回收率89.2%～100.6%,RSD2.2%～12.6%;AMOZ回收率91.3%～107.4%,RSD 4.3%～8.7%;SEM回收率79.9%～118.0%,RSD 2.4%～12.3%和A皿回收率78.9%～105.0%,RSD3.7%～12.1%.该快速检测方法小批量(20个样品)检测周期少于5.5h.     

反相高效液相色谱法检测虾青素

建立了一种快速检测虾青素的高效液相色谱法。色谱柱为D ikm a D iamonsilTM-C18(250 mm×4.6 mmi.d.,5μm),柱温为25℃,甲醇为流动相,检测波长为478 nm。虾青素的质量浓度在1~10μg/mL内与色谱峰面积线性关系良好,相关系数为0.999 7,该方法测定结果的相对标准偏差为0.38%(n=6),回收率为99.7%。     

虾青素简介

虾青素是一种极具发展潜力的抗氧化剂和色素,在医药、食品、水产养殖等方面具有广泛的应用。对虾青素的结构、性质、生理功能、应用、生产方法及发展前景等进行了简要介绍。     

Production and extraction of astaxanthin from Phaffia rhodozyma and its biological effect on alcohol-induced renal hypoxia in Carassius auratus

The effect of astaxanthin (3,3′-dihydroxy-s-carotene-4,4′-dione) on alcohol-induced morphological changes in Carassius auratus, as an experimental model, was determined. The yeast Phaffia rhodozyma was used as a source of astaxanthin. The animals were divided into three groups for 30 days: one group was treated with ethanol at a dose of 1.5% mixed in water, the second one with EtOH 1.5% and food enriched with astaxanthin from P. rhodozyma, and the third was a control group. After a sufficient experimental period, the samples were processed using light microscopy and evaluated by histomorphological and histochemical staining, and the data were supported by immunohistochemical analysis, using a wide range of antibodies, such as calbindin, vimentin and alpha-smooth muscle actin. The results show that the alcoholic damage in the kidney led to hypoxia. In contrast, the group fed with astaxanthin from P. rhodozyma showed a normal morphological picture, with better glomeruli organisation and the presence of the area of filtration. Furthermore, the immunohistochemistry has confirmed these results.     

Astaxanthin and its Effects in Inflammatory Responses and Inflammation-Associated Diseases: Recent Advances and Future Directions

Astaxanthin is a natural lipid-soluble and red-orange carotenoid. Due to its strong antioxidant property, anti-inflammatory, anti-apoptotic, and immune modulation, astaxanthin has gained growing interest as a multi-target pharmacological agent against various diseases. In the current review, the anti-inflammation mechanisms of astaxanthin involved in targeting for inflammatory biomarkers and multiple signaling pathways, including PI3K/AKT, Nrf2, NF-κB, ERK1/2, JNK, p38 MAPK, and JAK-2/STAT-3, have been described. Furthermore, the applications of anti-inflammatory effects of astaxanthin in neurological diseases, diabetes, gastrointestinal diseases, hepatic and renal diseases, eye and skin disorders, are highlighted. In addition to the protective effects of astaxanthin in various chronic and acute diseases, we also summarize recent advances for the inconsistent roles of astaxanthin in infectious diseases, and give our view that the exact function of astaxanthin in response to different pathogen infection and the potential protective effects of astaxanthin in viral infectious diseases should be important research directions in the future.     

Labeled vs. Label-Free Raman Imaging of Lipids in Endothelial Cells of Various Origins

Endothelial cells (EC) constitute a single layer of the lining of blood vessels and play an important role in maintaining cardiovascular homeostasis. Endothelial dysfunction has been recognized as a primary or secondary cause of many diseases and it manifests itself, among others, by increased lipid content or a change in the lipid composition in the EC. Therefore, the analysis of cellular lipids is crucial to understand the mechanisms of disease development. Tumor necrosis factor alpha (TNF-α)-induced inflammation of EC alters the lipid content of cells, which can be detected by Raman spectroscopy. By default, lipid detection is carried out in a label-free manner, and these compounds are recognized based on their spectral profile characteristics. We consider (3S,3′S)-astaxanthin (AXT), a natural dye with a characteristic resonance spectrum, as a new Raman probe for the detection of lipids in the EC of various vascular beds, i.e., the aorta, brain and heart. AXT colocalizes with lipids in cells, enabling imaging of lipid-rich cellular components in a time-dependent manner using laser power 10 times lower than that commonly used to measure biological samples. The results show that AXT can be used to study lipids distribution in EC at various locations, suggesting its use as a universal probe for studying cellular lipids using Raman spectroscopy. The use of labeled Raman imaging of lipids in the EC of various organs could contribute to their easier identification and to a better understanding of the development and progression of various vascular diseases, and it could also potentially improve their diagnosis and treatment.     

Effect of CO2 Flow Rate on the Extraction of Astaxanthin and Fatty Acids from Haematococcus pluvialis Using Supercritical Fluid Technology

Haematococcus pluvialis is the largest producer of natural astaxanthin in the world. Astaxanthin is a bioactive compound used in food, feed, nutraceutics, and cosmetics. In this study, astaxanthin extraction from H. pluvialis by supercritical fluid extraction was evaluated. The effects of temperature (40 and 50 °C), pressure (40 and 50 MPa), and CO₂ flow rate (2 and 4 L/min) were investigated. The results showed that the highest astaxanthin recovery was obtained at 50 °C/50 MPa and the CO₂ flow rates evaluated had no significant effect. It was possible to achieve astaxanthin recoveries of 95% after 175 min for a CO₂ flow rate of 2 L/min, and 95 min for CO₂ flow rate of 4 L/min. The ω-6/ω-3 ratios obtained were similar in all conditions, reaching 0.87, demonstrating that the extracts from H. pluvialis by SFE are rich in unsaturated fatty acids (UFA) which increases their positive effects when used as a functional ingredient in food.     

Two‐dimensional liquid chromatography analysis of all‐trans‐, 9‐cis‐, and 13‐cis‐astaxanthin in raw extracts from Phaffia rhodozyma

An effective two‐dimensional liquid chromatography method has been established for the analysis of all‐trans‐astaxanthin and its geometric isomers from Phaffia rhodozyma employing a C18 column at the first dimension and a C30 column in the second dimension, connected by a 10‐port valve using the photo‐diode array detector. The regression equation of astaxanthin calibration curve was established, and the precision and accuracy values were found to be in the range of 0.32–1.14% and 98.21–106.13%, respectively. By using two‐dimensional liquid chromatography, it was found that day light, ultrasonic treatment, and heat treatment have significant influence on the content of all‐trans‐astaxanthin in the extract from P. rhodozyma due to the transformation of all‐trans‐astaxanthin to cis‐astaxanthin. The day light and ultrasonic treatments more likely transform all‐trans‐astaxanthin to 9‐cis‐astaxanthin, and the thermal treatment transforms all‐trans‐astaxanthin to 13‐cis‐astaxanthin. These results indicate that the two‐dimensional liquid chromatography method can facilitate monitoring astaxanthin isomerization in the raw extract from P. rhodozyma. In addition, the study will provide a general reference for monitoring other medicals and bioactive chemicals with geometric isomers.     

HPLC Quantification of astaxanthin and canthaxanthin in Salmonidae eggs

Astaxanthin and canthaxanthin are naturally occurring antioxidants referred to as xanthophylls. They are used as food additives in fish farms to improve the organoleptic qualities of salmonid products and to prevent reproductive diseases. This study reports the development and single‐laboratory validation of a rapid method for quantification of astaxanthin and canthaxanthin in eggs of rainbow trout (Oncorhynchus mykiss ) and brook trout (Salvelinus fontinalis М.). An advantage of the proposed method is the perfect combination of selective extraction of the xanthophylls and analysis of the extract by high‐performance liquid chromatography and photodiode array detection. The method validation was carried out in terms of linearity, accuracy, precision, recovery and limits of detection and quantification. The method was applied for simultaneous quantification of the two xanthophylls in eggs of rainbow trout and brook trout after their selective extraction. The results show that astaxanthin accumulations in salmonid fish eggs are larger than those of canthaxanthin. As the levels of these two xanthophylls affect fish fertility, this method can be used to improve the nutritional quality and to minimize the occurrence of the M74 syndrome in fish populations.     

Determination of Astaxanthin and Canthaxanthin Triplet Properties in Different Polarities of the Solvent by Laser Flash Photolysis

Triplet‐triplet extinction coefficients for astaxanthin ( I ) and canthaxanthin ( II ) in different deaerated polarity solutions of MeCN and benzene were evaluated by laser flash photolysis at 298 K in the spectral region from 350 to 650 nm by energy transfer method, employing 2‐acetonaphthone as sensitizer. The triplet‐triplet extinction coefficients in MeCN and benzene were different in terms of the carotenoid present. The maximum triplet‐triplet extinction coefficient was 0.1–1.7×10⁵ L·mol⁻¹·cm⁻¹ in different solvents. The rate constants of triplet decay were I : 1.25×10¹⁰ L·mol⁻¹·s⁻¹, II : 1.12×10¹⁰ L·mol⁻¹·s⁻¹ in MeCN; and I : 1.75×10¹⁰ L·mol⁻¹·cm⁻¹, II : 3.27×10¹⁰ L·mol⁻¹·s⁻¹ in benzene. The bimolecular rate constants of energy transfer from triplet excited 2‐acetonaphthone to carotenoids were determined from the linear regression of the decay rate constant of 2‐acetonaphthone triplet at varying carotenoid concentrations. The triplet lifetimes of ³AST* and ³CAN* in different solvents were also determined. The results indicated that triplet energy transfer was nearly diffusion‐controlled.