Micellar casein concentrate production with a 3X, 3-stage, uniform transmembrane pressure ceramic membrane process at 50°C.

The production of serum protein (SP) and micellar casein from skim milk can be accomplished using microfiltration (MF). Potential commercial applications exist for both SP and micellar casein. Our research objective was to determine the total SP removal and SP removal for each stage, and the composition of retentates and permeates, for a 3×, continuous bleed-and-feed, 3-stage, uniform transmembrane pressure (UTP) system with 0.1-µm ceramic membranes, when processing pasteurized skim milk at 50°C with 2 stages of water diafiltration. For each of 4 replicates, about 1,100 kg of skim milk was pasteurized (72°C, 16s) and processed at 3× through the UTP MF system. Retentate from stage 1 was cooled to <4°C and stored until the next processing day, when it was diluted with reverse osmosis water back to a 1× concentration and again processed through the MF system (stage 2) to a 3× concentration. The retentate from stage 2 was stored at <4°C, and, on the next processing day, was diluted with reverse osmosis water back to a 1× concentration, before running through the MF system at 3× for a total of 3 stages. The retentate and permeate from each stage were analyzed for total nitrogen, noncasein nitrogen, and nonprotein nitrogen using Kjeldahl methods; sodium dodecyl sulfate-PAGE analysis was also performed on the retentates from each stage. Theoretically, a 3-stage, 3× MF process could remove 97% of the SP from skim milk, with a cumulative SP removal of 68 and 90% after the first and second stages, respectively. The cumulative SP removal using a 3-stage, 3× MF process with a UTP system with 0.01-µm ceramic membranes in this experiment was 64.8 ± 0.8, 87.8 ± 1.6, and 98.3 ± 2.3% for the first, second, and third stages, respectively, when calculated using the mass of SP removed in the permeate of each stage. Various methods of calculation of SP removal were evaluated. Given the analytical limitations in the various methods for measuring SP removal, calculation of SP removal based on the mass of SP in the skim milk (determined by Kjeldahl) and the mass SP present in all of the permeate produced by the process (determined by Kjeldahl) provided the best estimate of SP removal for an MF process.

Production efficiency of micellar casein concentrate using polymeric spiral-wound microfiltration membranes.

Most current research has focused on using ceramic microfiltration (MF) membranes for micellar casein concentrate production, but little research has focused on the use of polymeric spiral-wound (SW) MF membranes. A method for the production of a serum protein (SP)-reduced micellar casein concentrate using SW MF was compared with a ceramic MF membrane. Pasteurized (79°C, 18s) skim milk (1,100 kg) was microfiltered at 50°C [about 3 × concentration] using a 0.3-µm polyvinylidene fluoride spiral-wound membrane, bleed-and-feed, 3-stage process, using 2 diafiltration stages, where the retentate was diluted 1:2 with reverse osmosis water. Skim milk, permeate, and retentate were analyzed for SP content, and the reduction of SP from skim milk was determined. Theoretically, 68% of the SP content of skim milk can be removed using a single-stage 3× MF. If 2 subsequent water diafiltration stages are used, an additional 22% and 7% of the SP can be removed, respectively, giving a total SP removal of 97%. Removal of SP greater than 95% has been achieved using a 0.1-µm pore size ceramic uniform transmembrane pressure (UTP) MF membrane after a 3-stage MF with diafiltration process. One stage of MF plus 2 stages of diafiltration of 50°C skim milk using a polyvinylidene fluoride polymeric SW 0.3-µm membrane yielded a total SP reduction of only 70.3% (stages 1, 2, and 3: 38.6, 20.8, and 10.9%, respectively). The SP removal rate for the polymeric SW MF membrane was lower in all 3 stages of processing (stages 1, 2, and 3: 0.05, 0.04, and 0.03 kg/m(2) per hour, respectively) than that of the comparable ceramic UTP MF membrane (stages 1, 2, and 3: 0.30, 0.11, and 0.06 kg/m(2) per hour, respectively), indicating that SW MF is less efficient at removing SP from 50°C skim milk than the ceramic UTP system. To estimate the number of steps required for the SW system to reach 95% SP removal, the third-stage SP removal rate (27.4% of the starting material SP content) was used to extrapolate that an additional 5 water diafiltration stages would be necessary, for a total of 8 stages, to remove 95% of the SP from skim milk. The 8-plus stages necessary to remove >95% SP for the SW MF membrane would create more permeate and a lengthier process than required with ceramic membranes.

Comparison of composition and sensory properties of 80% whey protein and milk serum protein concentrates.

Milk serum protein concentrates (SPC) are proteins found in cheese whey that are removed directly from milk. Because SPC are not exposed to the cheese-making process, enzymatic or chemical reactions that can lead to off-flavors are reduced. The objectives of this study were to identify and compare the composition, flavor, and volatile components of 80% protein SPC and whey protein concentrates (WPC). Each pair of 80% SPC and WPC was manufactured from the same lot of milk and this was replicated 3 times. At each replication, spray-dried product from each protein source was collected. Commercial 80% WPC were also collected from several manufacturers for sensory and volatile analyses. A trained sensory panel documented the sensory profiles of the rehydrated powders. Volatile components were extracted by solid-phase microextraction and solvent extraction followed by solvent-assisted flavor evaporation with gas chromatography-mass spectrometry and gas chromatography-olfactometry. Consumer acceptance testing of acidified 6% protein beverages made with 80% SPC and WPC produced in the pilot plant and with WPC from commercial sources was conducted. The SPC was lower in fat and had a higher pH than the WPC produced in the pilot plant or commercial WPC. Few sensory differences were found between the rehydrated SPC and WPC manufactured in this study, but their flavor profiles were distinct from the flavor of rehydrated commercial WPC. The pilot-plant WPC had higher concentrations of lipid oxidation products compared with SPC, which may be related to the higher fat content of WPC. There was a large difference in appearance between 80% SPC and WPC: solutions of SPC were clear and those of WPC were opaque. Concentrations of lipid oxidation products in commercial WPC were generally higher than those in pilot-plant SPC or WPC. Sensory profiles of the peach-flavored protein beverage included cereal, free fatty acid, and soapy flavors and bitter taste in beverages made from pilot-plant products, whereas cardboard flavors were detected in those made with commercial WPC. Consumer liking scores for the beverages made with SPC were ranked highest or equally high with beverages made with WPC for aroma, appearance, and mouthfeel, but the beverages made with SPC had lower flavor and overall liking scores compared with beverages made with 3 of the 4 WPC.

Comparison of composition, sensory, and volatile components of thirty-four percent whey protein and milk serum protein concentrates.

The objectives of this study were to identify and compare the composition, flavor, and volatile components of serum protein concentrate (SPC) and whey protein concentrate (WPC) containing about 34% protein made from the same milk to each other and to commercial 34% WPC from 6 different factories. The SPC and WPC were manufactured in triplicate with each pair of serum and traditional whey protein manufactured from the same lot of milk. At each replication, SPC and WPC were spray dried (SD) and freeze dried (FD) to determine the effect of the heat used in spray drying on sensory properties. A trained sensory panel documented the sensory profiles of rehydrated SD or FD powders. Volatile components were extracted by solid-phase microextraction (SPME) and solvent extraction followed by solvent-assisted flavor evaporation (SAFE) with gas chromatography-mass spectrometry and gas chromatography-olfactometry. Whey protein concentrates had higher fat content, calcium, and glycomacropeptide content than SPC. Color differences (Hunter L, a, b) were not evident between SPC and WPC powders, but when rehydrated, SPC solutions were clear, whereas WPC solutions were cloudy. No consistent differences were documented in sensory profiles of SD and FD SPC and WPC. The SD WPC had low but distinct buttery (diacetyl) and cardboard flavors, whereas the SD SPC did not. Sensory profiles of both rehydrated SD products were bland and lower in overall aroma and cardboard flavor compared with the commercial WPC. Twenty-nine aroma impact compounds were identified in the SPC and WPC. Lipid and protein oxidation products were present in both products. The SPC and WPC manufactured in this study had lower total volatiles and lower concentrations of many lipid oxidation compounds when compared with commercial WPC. Our results suggest that when SPC and WPC are manufactured under controlled conditions in a similar manner from the same milk using the same ultrafiltration equipment, there are few sensory differences but distinct compositional and physical property differences that may influence functionality. Furthermore, flavor (sensory and instrumental) properties of both pilot-scale manufactured protein powders were different from commercial powders suggesting the role of other influencing factors (e.g., milk supply, processing equipment, sanitation).

Efficiency of serum protein removal from skim milk with ceramic and polymeric membranes at 50 degrees C.

Raw milk (2,710 kg) was separated at 4 degrees C, the skim milk was pasteurized (72 degrees C, 16 s), split into 3 batches, and microfiltered using pilot-scale ceramic uniform transmembrane pressure (UTP; Membralox model EP1940GL0.1microA, 0.1 microm alumina, Pall Corp., East Hills, NY), ceramic graded permeability (GP; Membralox model EP1940GL0.1microAGP1020, 0.1 microm alumina, Pall Corp.), and polymeric spiral-wound (SW; model FG7838-OS0x-S, 0.3 microm polyvinylidene fluoride, Parker-Hannifin, Process Advanced Filtration Division, Tell City, IN) membranes. There were differences in flux among ceramic UTP, ceramic GP, and polymeric SW microfiltration membranes (54.08, 71.79, and 16.21 kg/m2 per hour, respectively) when processing skim milk at 50 degrees C in a continuous bleed-and-feed 3x process. These differences in flux among the membranes would influence the amount of membrane surface area required to process a given volume of milk in a given time. Further work is needed to determine if these differences in flux are maintained over longer processing times. The true protein contents of the microfiltration permeates from UTP and GP membranes were higher than from SW membranes (0.57, 0.56, and 0.38%, respectively). Sodium-dodecyl-sulfate-PAGE gels for permeates revealed a higher casein proportion in GP and SW permeate than in UTP permeate, with the highest passage of casein through the GP membrane under the operational conditions used in this study. The slight cloudiness of the permeates produced using the GP and SW systems may have been due to the presence of a small amount of casein, which may present an obstacle in their use in applications when clarity is an important functional characteristic. More beta-lactoglobulin passed through the ceramic membranes than through the polymeric membrane. The efficiency of removal of serum proteins in a continuous bleed-and-feed 3x process at 50 degrees C was 64.40% for UTP, 61.04% for GP, and 38.62% for SW microfiltration membranes. The SW polymeric membranes had a much higher rejection of serum proteins than did the ceramic membranes, consistent with the sodium-dodecyl-sulfate PAGE data. Multiple stages and diafiltration would be required to produce a 60 to 65% serum protein reduced micellar casein concentrate with SW membranes, whereas only one stage would be needed for the ceramic membranes used in this study.

Telomerase activity and patient survival after surgery for gastric and oesophageal cancer.

OBJECTIVE: The ribonucleoprotein telomerase extends telomeres in cancer cells and has been proposed as a prognostic marker for cancer. We measured telomerase expression in proximal adenocarcinomas (those arising in the distal oesophagus or at the gastro-oesophageal junction) and distal adenocarcinomas (those arising in the corpus or antrum of the stomach) of the foregut, and correlated telomerase activity with pathological stage and post-operative survival. DESIGN: Surgical specimens were collected from patients undergoing resections for gastric and oesophageal carcinomas. Haematoxylin and eosin histology provided data on the pathological tumour stage and pathological node stage. METHODS: The telomerase activity of cancer specimens was determined using the telomeric repeat amplification protocol. A single pathologist, blinded to the results of the telomerase assays, reviewed all slides of cancers to assign T and N stages. RESULTS: The cancers exhibited a wide range of telomerase expression. There was no significant difference between the telomerase activity of proximal adenocarcinomas (median, 551 U; 95% confidence interval, 154-2394 U; n = 26) and distal adenocarcinomas (median, 703 U; 95% confidence interval, 139-1618 U; n = 20). Distal adenocarcinomas expressing high telomerase activity (greater than the median) were significantly more advanced with regard to T stage than distal cancers expressing low telomerase levels (less than the median; P = 0.03, Mann-Whitney test). In distal adenocarcinomas, high telomerase activity was associated with poor patient survival (median 3 months) compared to low telomerase activity (median survival 22.4 months; P = 0.01, log-rank test). No such differences were observed for proximal adenocarcinomas. CONCLUSIONS: There is a difference between gastric and oesophageal/gastro-oesophageal junction adenocarcinomas in terms of the relationship with telomerase expression and clinico-pathological variables. Among patients with distal gastric adenocarcinoma, telomerase activity correlates with markers of advanced disease, whereas this relationship does not hold true in oesophageal/gastro-oesophageal junction adenocarcinomas. Telomerase activation may occur at different stages of the formation of the malignant phenotype in these two cancers and may reflect differences in their pathogenesis. Telomerase could be a prognostic marker in gastric but not in oesophageal adenocarcinoma.

Deficiency of colonic telomerase in ulcerative colitis.

OBJECTIVE: GI epithelial cells express telomerase, a ribonucleoprotein that prevents telomeric shortening in proliferating cells. Telomerase levels are high in cancer, but little is known about telomerase expression in other diseases. We, therefore, designed experiments to determine telomerase expression in different colonic segments and to compare this with corresponding segments in patients with ulcerative colitis. Colorectal cancers and adenomatous polyps were included as disease controls. METHODS: In total, telomerase expression was determined in colonic tissues obtained from 62 patients. Twenty-five patients had ulcerative colitis, 21 had normal colons, 11 had colorectal cancer, and nine had adenomatous polyps. Endoscopic biopsies were collected prospectively at colonoscopy, processed for telomerase assays (Telomeric Repeat Amplification Protocol), hematoxylin and eosin staining, and scored for inflammation. RESULTS: Telomerase activity is expressed in arbitrary units (median 95% confidence interval). In the normal colon, telomerase activity in the cecum, transverse, sigmoid, and rectum was 255 (171-449), 707 (374-895), 561 (468-1426), and 563 (402-846), respectively. Telomerase was higher in the distal three segments when compared with the cecum (p = 0.005). In ulcerative colitis, there was a marked decrease in telomerase activity in the cecum 152 (59-272), p = 0.04, transverse 180 (129-365), p < 0.001, sigmoid 352 (114-464), p = 0.005, and rectum 180 (70-337), p = 0.001 when compared with normals. Telomerase activity correlated negatively with inflammation (r = -0.32, p = 0.001) and was also decreased in microscopically normal areas. Cancers expressed high levels of telomerase. CONCLUSIONS: Colonic mucosal expression of telomerase is reduced in ulcerative colitis. Levels are low even in microscopically normal mucosa, suggesting that telomerase deficiency may contribute to the pathogenesis of the disease.

The performance of volume selection sequences for in vivo NMR spectroscopy: implications for quantitative MRS.

Previous work has demonstrated that deficiencies in volume selection sequences used in magnetic resonance spectroscopy may compromise the quality of the spectra obtained. In this paper, further studies on the ISIS and PRESS sequences are presented. Under conditions of partial saturation, ISIS can exhibit serious contamination with extraneous signal, particularly when a small volume of interest (VOI) is selected. ISIS protocols should therefore use VOIs that are large relative to the target volume, and repetition times that are as long as practicable. In PRESS, contamination is found to be minimised by using a VOI that is small relative to the target volume, and to be independent of repetition time. PRESS performance is also independent of echo time, except when very short echo times are used. These results are consistent with previously published work on ISIS and PRESS, and it is now possible to establish generic features of these sequences and to understand the implications for quantitative spectroscopy. T(1)-weighting of contamination in ISIS can compromise both relative and absolute quantification techniques in several respects. Contamination in PRESS is largely independent of relaxation times and would be easier to model and correct for in the context of quantitative spectroscopy.

Absolute metabolite quantification by in vivo NMR spectroscopy: II. A multicentre trial of protocols for in vivo localised proton studies of human brain.

We have performed a multicentre trial to assess the performance of three techniques for absolute quantification of cerebral metabolites using in vivo proton nuclear magnetic resonance (NMR). The techniques included were 1) an internal water standard method, 2) an external standard method based on phantom replacement, and 3) a more sophisticated method incorporating elements of both the internal and external standard approaches, together with compartmental analysis of brain water. Only the internal water standard technique could be readily implemented at all participating sites and gave acceptable precision and interlaboratory reproducibility. This method was insensitive to many of the experimental factors affecting the performance of the alternative techniques, including effects related to loading, standing waves and B1 inhomogeneities; and practical issues of phantom positioning, user expertise and examination duration. However, the internal water standard method assumes a value for the concentration of NMR-visible water within the spectroscopic volume of interest. In general, it is necessary to modify this assumed concentration on the basis of the grey matter, white matter and cerebrospinal fluid (CSF) content of the volume, and the NMR-visible water content of the grey and white matter fractions. Combining data from 11 sites, the concentrations of the principal NMR-visible metabolites in the brains of healthy subjects (age range 20-35 years) determined using the internal water standard method were (mean+/-SD): [NAA]=10.0+/-3.4 mM (n=53), [tCho]=1.9+/-1.0 mM (n=51), [Cr + PCr]=6.5+/-3.7 mM (n=51). Evidence of system instability and other sources of error at some participating sites reinforces the need for rigorous quality assurance in quantitative spectroscopy.

Neuroimaging and spectroscopy in children with epileptic encephalopathies.

AIMS: To investigate the nature of the unifocal cortical abnormalities on FDG positron emission tomography (PET) in children with an epileptic encephalopathy but no focal abnormality on electroencephalogram or standard magnetic resonance imaging (MRI). METHODS: Repeat FDG PET, surface rendered high resolution MRI, and single voxel magnetic resonance proton spectroscopy of the areas of abnormal metabolism compared to the contralateral side in 11 children aged 2 to 12 years. Imaging was repeated after a median of 13 months. RESULTS: Visual analysis of repeat FDG PET revealed similar abnormalities in 10 of 11 children. Semiquantitative analysis revealed similar sited abnormalities in eight children. One child with ictal hypermetabolism initially had an inconsistent second scan. Magnetic resonance spectra in three children showed the N-acetyl-aspartate/choline ratio was lower in the hypometabolic focus than in the reciprocal area on the opposite side, in two children it was higher, and in one child it was equal. Surface rendered MRI was normal in seven of eight children, and showed temporal lobe asymmetry in one. CONCLUSION: In children with established epileptic encephalopathies most hypometabolic areas on FDG PET are stable over time. While focal neuronal loss is likely in these areas in some children, microdysplasias or other focal cortical dysplasias are probable in others.

Preterm prelabour amniorrhexis: relation to histological chorioamnionitis.

Infection-mediated inflammation may be responsible for premature delivery in cases of preterm prelabour amniorrhexis. Histological examination of the cord and placenta was performed in 54 such pregnancies, in addition to amniocentesis and cordocentesis at presentation. Histological chorioamnionitis was more likely in those with positive amniotic fluid and/or fetal blood cultures, and in those cases with histological evidence of inflammation, the interval to delivery was short regardless of the culture results. Inflammation rather than infection per se may be the best indicator of preterm delivery in preterm prelabour amniorrhexis.

Pseudoreflux syndrome-increased periodic breathing during the neonatal period presenting as feeding-related difficulties.

Sixteen infants who presented with symptoms suggestive of gastroesophageal reflux (GER)-associated apnea were evaluated at the Breathing Disorder Center of the Children's Hospital of Philadelphia. These neonates had a history of occasional emesis and an apparent life-threatening event (ALTE) that occurred while awake which was similar to the presentation of a group of infants previously described. Evaluation of the present group of infants however, revealed increased periodic breathing (12.1 +/- 1.8 SEM% of total sleep time) as opposed to the obstructive apnea that was typically seen with GER. Pathologic gastroesophageal reflux could not be diagnosed in relationship to apneic events. Infants who present during the first month of life with symptoms suggestive of GER-associated apnea should have careful evaluation of reflux and respiratory patterns to confirm the correct diagnosis. Because of the similarities of these infants to the GER group, we have called their disorder pseudoreflux.

Reduced tumour growth of the human colonic cancer cell lines COLO-320 and HT-29 in vivo by dietary n-3 lipids.

Seventy-five nude mice received subcutaneous inoculation with 1 X 10(7) cells of the human colonic cancer cell lines COLO-320 or HT-29. Tumour growth was assessed over 4 weeks in animals given one of three iso-caloric diets; standard diet, high saturated fat (20% coconut) diet and high n-3 fat (20% Maxepa fish oil) diet. The n-3 diet produced significant tumour growth reduction compared to the other diets for COLO-320 at 3 to 4 weeks (P less than 0.05 at least) and similarly for HT-29 at 4 weeks (P less than 0.05). Significant incorporation of n-3 fatty acids occurred in red cell membranes, adipose tissue and both neutral lipid and phospholipid fractions of tumour lipids in animals fed Maxepa (P less than 0.01 at least). This was accompanied by reduction of linoleic acid and arachidonic acid in these tissues (P less than 0.01 at least) but was most marked in the metabolically labile phospholipid fraction. There was high mitotic activity in the tumours from all the groups but there was no difference according to diet.