Determination of bixin and norbixin in meat using liquid chromatography and photodiode array detection

The development of an analytical method that enables routine analysis of annatto dye, specifically bixin and norbixin, in meat tissue is described. Liquid-solid extraction was carried out using acetonitrile. Analysis was by HPLC with photodiode array detection using two fixed wavelengths (458 and 486 nm). The possibilities of ion trap mass spectrometry (MS) were also assessed. Method performance characteristics, according to Commission Decision 2002/657/EC, were determined, with recoveries between 99 and 102% and calibration curves being linear in the 0.5–10 mg kg^?1 range. The limit of quantification was 0.5 mg kg^?1.     

Short communication: Development of a novel method for the extraction of norbixin from whey and its subsequent quantification via high performance liquid chromatography

Norbixin is the primary carotenoid in annatto coloring, which imparts the desired orange color in Cheddar cheese. However, a portion of the colorant remains in the cheese whey and is undesirable; therefore, a bleaching step is often applied. Restrictions exist for norbixin concentrations in products destined for infant formula. As such, evaluation of norbixin concentrations in whey and whey ingredients is desirable. Current extraction methods are laborious and require solvents that are banned in many countries. The objective of this study was to develop a fast and inexpensive norbixin extraction and quantitation technique using approved solvents with similar sensitivity to current established methods. Instead of solvent extraction and column purification, acetonitrile was added directly to fluid wheys, retentates, and rehydrated whey protein concentrates. An isocratic mobile phase [70% acetonitrile and 30% water with 0.1% (wt/vol) formic acid] was used and, to increase sensitivity, a large volume (50 μL) was injected onto the column. The column used was a C18 column with a particle size of 2.6 μm and column length of 10 cm. The column inner diameter was 4.6 mm and the pore size was 100 ?. All of the previously described conditions allowed the run time to be only 4 min. The sample was sent through a photodiode array detector and quantified at 482 nm. Norbixin was quantified using external standard curves. The developed method had a >90% norbixin recovery in both milk and whey (9.39 μg/L–2.35 mg/L). The limit of detection of norbixin in fluid whey was 2.7 μg/kg and the limit of quantitation was 3.5 μg/kg, both of which are significantly lower than in previously described methods. The extracts were stable over 30 min at 21°C and stable over 24 h at 4°C. Repeatability and precision of the method had relative standard deviations of less than 13%. The developed method provides time and cost savings for evaluation of norbixin concentration in whey and whey products.     

Colorants in cheese manufacture: Production,chemistry, interactions,and regulation

Colored Cheddar cheeses are prepared by adding an aqueous annatto extract (norbixin) to cheese milk; however, a considerable proportion (～20%) of such colorant is transferred to whey, which can limit the end use applications of whey products. Different geographical regions have adopted various strategies for handling whey derived from colored cheeses production. For example, in the United States, whey products are treated with oxidizing agents such as hydrogen peroxide and benzoyl peroxide to obtain white and colorless spray‐dried products; however, chemical bleaching of whey is prohibited in Europe and China. Fundamental studies have focused on understanding the interactions between colorants molecules and various components of cheese. In addition, the selective delivery of colorants to the cheese curd through approaches such as encapsulated norbixin and microcapsules of bixin or use of alternative colorants, including fat‐soluble/emulsified versions of annatto or beta‐carotene, has been studied. This review provides a critical analysis of pertinent scientific and patent literature pertaining to colorant delivery in cheese and various types of colorant products on the market for cheese manufacture, and also considers interactions between colorant molecules and cheese components; various strategies for elimination of color transfer to whey during cheese manufacture are also discussed.     

Reflectance versus transmittance: The effects of light scattering on red colorants (carmine,amazonian red annatto,and peruvian cochinilla rojo and rosada) in biological,textile, and museum science

Spectra were strongly influenced by the way they were measured but, in general, carmine, cochinilla rojo and rosada, and red annatto all had strong transmittance and reflectance of red light (>640 nm), and strong absorbance of green light (520–560 nm). Spectra for carmine and cochinilla rosada both had secondary peaks for transmittance and reflectance (around 420 and 450 nm for stains, respectively), whereas secondary peaks were not found for cochinilla rojo and red annatto. Both cochinilla rosada and rojo stained skeletal myofibers, but only cochinilla rojo withstood alcohol dehydration and mounting as a general stain for paraffin sections. Carmine was ideal for enhancing the appearance of pork because its spectrum was similar to that of myoglobin, thus increasing the absorbance of green light without producing unnatural colors like cochinilla rosada and rojo. Cochinilla rojo dyed alpaca wool orange, and cochinilla rosada dyed wool pink. The scale of the measurement (micro‐ vs. macroscopic) and the type of measurement (transmittance vs. reflectance) were of minor importance in colorimetry, whereas a major effect was detected for light scattering in the sample. A ratio indicative of scattering (400/700 nm) was strongly correlated with chromaticity coordinate x, r = ?0.86, P < 0.001, n = 18. © 2013 Wiley Periodicals, Inc. Col Res Appl, 39, 599–606, 2014     

BIXIN POWDER PRODUCTION IN CONICAL SPOUTED BED UNITS

ABSTRACT

Recent work has demonstrated that the red carotenoid bixin is easily extracted from Bixa orellana seeds by particle attrition and impact rather than by solvent extraction. This technique may require a previous step for drying seeds at a safe temperature to preserve pigment quality. A spouted bed (SB) with a draft tube is the most appropriate equipment to process both seed drying and bixin extraction at low operational costs. Operational parameters of the SB unit are optimized as a function of the powder production rate. The maximum air inlet temperature for drying the seeds is also specified. A high performance liquid chromatography method has been developed for the semi-quantitative determination of bixin content in the powder produced. Results show that this SB unit can be more competitive than ball mill equipment.     

高效液相色谱法测定巧克力中胭脂树橙的含量

目的建立高效液相色谱法测定巧克力中胭脂树橙含量的分析方法。方法采用氨水乙醇提取样品中的色素,石油醚除脂后,在酸性条件下采用二氯甲烷反提取目标物,选用C_(18)色谱柱分离,以乙腈:2%乙酸(75:25,V:V)作为流动相进行等度洗脱,用紫外检测器在458 nm波长检测,外标法定量。结果降红木素的定量限为1.0 mg/kg,红木素的定量限为2.0 mg/kg,线性范围为1～50 mg/kg。降红木素和红木素回收率分别为87.8%～92.0%和88.7%～92.1%,相对标准偏差(relative standard deviation,RSD)分别为1.1%～5.0%和1.7%～5.2%。结论本方法简单、便捷,适用于巧克力中胭脂树橙含量的测定。     

Effect of Storage on the Stability of Water Soluble Annatto Dye Formulation in a Simulated Orange-RTS Beverage Model System

Annatto (Bixa orellana L.) seeds are the source of an orange yellow pigment. The water-soluble annatto dye formulations can be used in beverages, bakery and other food products. An annatto water-soluble dye formulation containing norbixin fraction 22.4 g/L was used in making an orange-ready-to-serve (RTS) beverage model with citric acid and sugar. The orange-RTS beverage was stored at ambient and refrigerated conditions in transparent as well as amber-coloured glass bottles for a period of 5 mo. A dilute solution of the annatto dye formulation (working stock) without addition of citric acid and sugar was also stored under similar conditions. The annatto water-soluble dye formulation and the orange-RTS beverage stored at refrigerated conditions stored well, whereas the losses were very high for the working stock of the formulation irrespective of the storage period/method.     

Tiered intake assessment for food colours

ABSTRACT

A tiered intake assessment approach, ranging from the conservative default and refined budget method to refined dietary exposure assessments using national food consumption surveys is presented and applied to derive maximum potential global colour intake estimates. The US and UK markets served as representative for the world and the EU, respectively, to determine the maximum potential exposure ceilings for eleven colours in various sub-populations, including brand-loyal consumers. Industry-reported global use levels were assigned as the maximum level. Conservative intake assessments for food colours used in non-alcoholic beverages were estimated for the general population 2 + y, toddlers, children 3–9 y, adolescents 10–17 y, adults 18–64 y, elderly 65–74 y, very elderly 75 + y based on assumed uses in high intake markets. Refined dietary exposures were estimated using either the 2-day food consumption data from the 2013–2016 US National Health and Nutrition Examination Survey or the 4-day food consumption data from the 2008–2016 U.K. National Diet and Nutrition Survey Rolling Programme. In the most refined market-share adjusted assessment, brand-specific market volume data were used to place appropriate weight on corresponding beverage type uses. Strong concordance between the refined budget method and the brand-loyal deterministic approach was shown, in which the latter assumes that the maximum use level of the colour is present in 100% of non-alcoholic beverages. This study shows that safety of colours – both synthetic and natural – in beverages at proposed use levels can be supported for any geography, with all intake estimates falling below the acceptable daily intake in refined assessments. Importantly, this study demonstrates that the refined budget method is a valid first-tier screening assessment to prioritise food colours that may benefit from more refined intake assessments when warranted.     

The effect of starter culture and annatto on the flavor and functionality of whey protein concentrate

The flavor of whey protein can carry over into ingredient applications and negatively influence consumer acceptance. Understanding sources of flavors in whey protein is crucial to minimize flavor. The objective of this study was to evaluate the effect of annatto color and starter culture on the flavor and functionality of whey protein concentrate (WPC). Cheddar cheese whey with and without annatto (15 mL of annatto/454 kg of milk, annatto with 3% wt/vol norbixin content) was manufactured using a mesophilic lactic starter culture or by addition of lactic acid and rennet (rennet set). Pasteurized fat-separated whey was then ultrafiltered and spray dried into WPC. The experiment was replicated 4 times. Flavor of liquid wheys and WPC were evaluated by sensory and instrumental volatile analyses. In addition to flavor evaluations on WPC, color analysis (Hunter Lab and norbixin extraction) and functionality tests (solubility and heat stability) also were performed. Both main effects (annatto, starter) and interactions were investigated. No differences in sensory properties or functionality were observed among WPC. Lipid oxidation compounds were higher in WPC manufactured from whey with starter culture compared with WPC from rennet-set whey. The WPC with annatto had higher concentrations of p-xylene, diacetyl, pentanal, and decanal compared with WPC without annatto. Interactions were observed between starter and annatto for hexanal, suggesting that annatto may have an antioxidant effect when present in whey made with starter culture. Results suggest that annatto has a no effect on whey protein flavor, but that the starter culture has a large influence on the oxidative stability of whey.     

BIXIN POWDER PRODUCTION IN CONICAL SPOUTED BED UNITS

Recent work has demonstrated that the red carotenoid bixin is easily extracted from Bixa orellana seeds by particle attrition and impact rather than by solvent extraction. This technique may require a previous step for drying seeds at a safe temperature to preserve pigment quality. A spouted bed (SB) with a draft tube is the most appropriate equipment to process both seed drying and bixin extraction at low operational costs. Operational parameters of the SB unit are optimized as a function of the powder production rate. The maximum air inlet temperature for drying the seeds is also specified. A high performance liquid chromatography method has been developed for the semi-quantitative determination of bixin content in the powder produced. Results show that this SB unit can be more competitive than ball mill equipment.