Rapid Isoelectric Point Determination in a Miniaturized Preparative Separation Using Jet-Dispensed Optical pH Sensors and Micro Free-Flow Electrophoresis

Herein, the fabrication, characterization, calibration, and application of integrated microfluidic platforms for fast isoelectric point (pI) determinations via free-flow electrophoresis with integrated inkjet-printed fluorescent pH sensor microstructures are presented. These devices allow one to determine the pI of a biomolecule from a sample mixture with moderately good precision and without addition of markers in typically less than 10 s total separation and analysis time. Polyhydroxyethyl methacrylate (pHEMA) hydrogels were covalently coupled with fluorescein and hydroxypyrene trisulfonic acid (HPTS)-based pH probes. These were piezoelectrically jet-dispensed onto acrylate-modified glass as pH sensor microarrays with a diameter of 300–600 μm and thicknesses of 0.4–2.4 μm with high spatial accuracy. Microchip fabrication and integration of these pH sensor arrays was realized by multistep liquid-phase photolithography from oligoethylene glycol precursors resulting in glass-based microfluidic free-flow isoelectric focusing (μFFIEF) chips with integrated pH observation capabilities. The microchips were characterized with regard to pH sensitivity, response times, photo-, and flow stability. Depending on the sensor matrix, they allowed IEF within a pH range of roughly 5.5–10.5 with good sensitivity and fast response times. These microchips were used for FFIEF of small molecule markers and several protein mixtures with simultaneous monitoring of local pH. This allowed the determination of their pI via multispectral imaging of protein and pH sensor fluorescence without addition of external markers. Obtained pI’s were generally in good agreement with known data, demonstrating the applicability of the method for pI determination in micropreparative procedures within a time frame of a few seconds only

Mechanisms Regulating Insulin Response to Intragastric Glucose in Lean and Non-Diabetic Obese Subjects: A Randomized, Double-Blind, Parallel-Group Trial

BACKGROUND/OBJECTIVES: The changes in blood glucose concentrations that result from an oral glucose challenge are dependent on the rate of gastric emptying, the rate of glucose absorption and the rate of insulin-driven metabolism that include the incretins, glucose-dependent insulinotropic peptide (GIP) and glucagon-like peptide-1 (GLP-1). The rate of insulin-driven metabolism is clearly altered in obese subjects, but it is controversial which of these factors is predominant. We aimed to quantify gastric emptying, plasma insulin, C-peptide, glucagon and glucose responses, as well as incretin hormone secretions in obese subjects and healthy controls during increasing glucose loads. SUBJECTS/METHODS: The study was conducted as a randomized, double-blind, parallel-group trial in a hospital research unit. A total of 12 normal weight (6 men and 6 women) and 12 non-diabetic obese (BMI > 30, 6 men and 6 women) participants took part in the study. Subjects received intragastric loads of 10 g, 25 g and 75 g glucose dissolved in 300 ml tap water. RESULTS: Main outcome measures were plasma GLP-1 and GIP, plasma glucagon, glucose, insulin, C-peptide and gastric emptying. The primary findings are: i) insulin resistance (P < 0.001) and hyperinsulinemia (P < 0.001); ii) decreased insulin disposal (P < 0.001); iii) trend for reduced GLP-1 responses at 75 g glucose; and iv) increased fasting glucagon levels (P < 0.001) in obese subjects. CONCLUSIONS: It seems that, rather than changes in incretin secretion, fasting hyperglucagonemia and consequent hyperglycemia play a role in reduced disposal of insulin, contributing to hyperinsulinemia and insulin resistance. TRIAL REGISTRATION: ClinicalTrials.gov NCT01875575.status: publishe

Migration of aluminum from food contact materials to food—a health risk for consumers? Part II of III: migration of aluminum from drinking bottles and moka pots made of aluminum to beverages

Abstract Background Drinking bottles and stove-top moka pots made of aluminum have become very popular. Storing drinks in bottles and preparing coffee in a moka pot may result in the migration of aluminum to the beverage. Results/Conclusions In a systematic study of aluminum drinking bottles, it has been shown that drinking a mixture of apple juice and mineral water in an aluminum bottle may reach 86.6% of the total weekly intake (TWI) for adults, and drinking tea from an aluminum bottle may exceed the TWI (145%) for a child weighing 15 kg. In contrast, preparing coffee in an aluminum moka pot results in a maximum of 4% to TWI, if an average of 3.17 L coffee is consumed per week, even if the pots are washed in the dishwasher, against the explicit instructions of the manufacturer

Migration of aluminum from food contact materials to food—a health risk for consumers? Part I of III: exposure to aluminum, release of aluminum, tolerable weekly intake (TWI), toxicological effects of aluminum, study design, and methods

Abstract Background In spite of the prevalence of aluminum in nature, no organism has been found to date which requires this element for its biological functions. The possible health risks to human beings resulting from uptake of aluminum include detrimental effects to the hemopoietic system, the nervous system and bones. Aluminum is used in many fields and occurs in numerous foodstuffs. Food contact materials containing aluminum represent an anthropogenic source of dietary aluminum. Results As a result of their frequent use in private households a study was undertaken to detect migration of this metal to foodstuffs from drink containers, coffee pots, grill pans, and camping cookware made of aluminum. Conclusions An estimate of the health risk to consumers is calculated, based on the tolerable weekly intake (TWI) specified by the European Food Safety Authority of 1 mg/kg body weight for all groups of people. In some instances the TWI is significantly exceeded, dependent upon the food contact material and the food itself

Migration of aluminum from food contact materials to food—a health risk for consumers? Part III of III: migration of aluminum to food from camping dishes and utensils made of aluminum

Abstract Background When cooking on a barbecue grill, consumers often use aluminum grill pans. For one, the pan catches the fats and oils that would drip into the embers causing the formation of potentially noxious smoke, and the pan also protects the food from being burned by direct heat from the coals. In addition, new aluminum products for use in ovens and grills are becoming increasingly popular. Due to their light weight and excellent heat transfer camping, utensils made of aluminum are, for example, often used by fishermen and mountain climbers. Preparing food in aluminum utensils can, however, result in migration of the aluminum to the foodstuffs. Results/Conclusions In this study presented here, it was found that the transfer limit of 5.00 mg/L for aluminum is not exceeded using simulants for oil or for tap water; however, with an aqueous solution of 0.5% citric acid, the limit is clearly exceeded at 638 mg/L. This means that the Tolerable Weekly Intake (TWI) is exceeded by 298% for a child weighing 15 kg and for an adult weighing 70 kg it is equivalent to 63.8% of the TWI, assuming a daily uptake of 10 mL marinade containing lemon juice over a period of 1 week. Preparation of a fish dish with a marinade containing lemon juice in camping dishes would result in the TWI being exceeded by 871% for a child weighing 15 kg and by 187% for an adult weighing 70 kg assuming a daily uptake of 250 g over a period of 1 week

Graphical genotypes of introgression populations resulting from six different crossing schemes.

<p>A: ; B: ; C: ; D: ; E: ; F: . The graphical genotypes display the chromosomes 3 to 7 of ILs 41–70 and are examples from one simulation run. Chromosome segments which stem from the donor are displayed in blue, whereas chromosome segments which stem from the recipient are displayed in yellow. The graphical genotypes illustrate the differences between the alternative crossing schemes with respect to their suitability to create introgression populations with complete donor genome coverage and clearly separated, evenly distributed target donor chromosome segments.</p

Measures evaluated for introgression populations resulting from different crossing and selection schemes.

<p>: donor genome coverage in percent; : depth of donor genome coverage; : number of disjunct genome segments; : resolution; : number of donor segments per IL; : length of donor segments per IL in cM; : total donor genome proportion in percent; : donor genome proportion of carrier chromosomes in percent; : donor genome proportion of target segments in percent; HT: the required number of HT assays. Measures are arithmetic means over 1,000 replications.</p

Schematic representation of crossing scheme .

<p>Crossing scheme is characterized by increasing population sizes in the backcross generations and selection for donor chromosome halves in the final backcross generation. The parts highlighted in gray represent one branch of the crossing scheme. Sub-populations are indexed by , and , where is the respective backcross generation, is the respective chromosome, is the respective chromosome half, is the respective target segment; and denote individuals selected for the respective selection regions.</p

Definition of the selection index in generations , , , DH, , for different selection strategies for developing introgression populations.

<p>Selection for complete donor chromosomes (C), selection for donor chromosome halves (H) and selection for donor target segments (S) were combined to form different selection strategies (left column). , and denote the donor genome proportions of the chromosome on which the target segment is located, of the chromosome half on which the target segment is located and of the target segment itself. , and correspond to , , and denote the recipient genome proportion outside the respective chromosome region.</p