A targeted GC-MS/MS approach for the determination of eight sterols in microgreen and mature plant material.

Over the past decades, a remarkable number of scientific studies supported the correlation between an adequate dietary intake of phytosterols (PS) and the reduced risk of cardiovascular diseases. PS are known to inhibit the intestinal absorption of cholesterol, thus promoting the reduction of the low-density lipoproteins (LDL) amount in the bloodstream. Despite the fact that a non-negligible atherogenicity was recognized to PS, thus requiring a careful risk-benefits assessment for plant sterol supplementation, the potential role of PS as cholesterol-lowering agents has been contributing to the spreading awareness of the health benefits associated with the consumption of plant-based foods. In recent years, this has been fueling the market of innovative vegetable products, such as microgreens. Surprisingly, the recent literature concerning microgreens exhibited the lack of studies focusing on the characterization of PS. To fill this gap, a validated analytical method based on the hyphenation of gas chromatography and tandem mass spectrometry is proposed here for the quantitative analysis of eight phytosterols, namely ß-sitosterol, campesterol, stigmasterol, brassicasterol, isofucosterol, and cholesterol, lathosterol and lanosterol. The method was exploited for the characterization of the PS content in 10 microgreen crops, i.e., chia, flax, soybean, sunflower, rapeseed, garden cress, catalogna chicory, endive, kale and broccoli raab. Finally, these results were compared to the PS content of mature forms of kale and broccoli raab. A remarkable amount of PS was detected in chia, flax, rapeseed, garden cress, kale, and broccoli raab microgreens. 100 g (wet weight) of these microgreen crops were found to contain from 20 to 30 mg of the investigated PS. Interestingly, in the case of kale and broccoli raab microgreens, the overall PS content was higher than the one measured in the edible parts of the corresponding mature forms. Additionally, a symmetric change of the PS inner profile was observed between the two growth stages of the latter two crops. Here, the overall decrease of the PS sterol content in the mature forms was associated with the increase of the relative amount of ß-sitosterol and campesterol at the expense of minor PS species, such as brassicasterol.

Insight into the Storage-Related Oxidative/Hydrolytic Degradation of Olive Oil Secoiridoids by Liquid Chromatography and High-Resolution Fourier Transform Mass Spectrometry.

The study of negative effects potentially exerted by the exposure to oxygen and/or light and, thus, also by the type of container on the quality of extra virgin olive oil (EVOO) during its prolonged storage requires an appropriate choice of analytical methods and components to be monitored. Here, reverse-phase liquid chromatography coupled to high-resolution/accuracy Fourier transform mass spectrometry with electrospray ionization was exploited to study oxidative/hydrolytic degradation processes occurring on the important bioactive components of EVOO known as secoiridoids, i.e., oleuropein and ligstroside aglycones, oleacin, and oleocanthal, during storage up to 6 months under controlled conditions. Specifically, isomeric oxidative byproducts resulting from the transformation of a carbonylic group of the original secoiridoids into a carboxylic group and compounds resulting from hydrolysis of the ester linkage of secoiridoids, i.e., elenolic and decarboxymethyl elenolic acids and tyrosol and 3-hydroxytyrosol, were monitored, along with their precursors. Data obtained from EVOO storage at room temperature in glass bottles with/without exposure to light and/or oxygen indicated that, although it was more relevant if a periodical exposure to oxygen was performed, a non-negligible oxidative degradation occurred on secoiridoids also when nitrogen was used to saturate the container headspace. In a parallel experiment, the effects of storage of the same EVOO (250 mL) for up to 6 months in containers manufactured with different materials/shapes were considered. In particular, a square dark glass bottle, a stainless-steel can, and a ceramic jar, typically used for EVOO commercialization, and a clear polyethylene terephthalate bottle, purposely chosen to prompt secoiridoid degradation through exposure to light and oxygen, were compared. Dark glass was found to provide the best combined protection of major secoiridoids from oxidative and hydrolytic degradation, yet the lowest levels of oxidized byproducts were observed when the stainless-steel can was used.

The occurrence of inositolphosphoceramides in spirulina microalgae.

Spirulina microalga (Arthrospira platensis) is an interesting phototrophic organism because of its high content of nutrients including proteins, lipids, essential amino acids, antioxidants, vitamins, polysaccharides, and minerals. Hydrophilic interaction liquid chromatography (HILIC) coupled to linear ion trap (LIT) and Orbitrap Fourier transform mass spectrometry (FTMS) via ESI was employed for the separation and characterization of lipid species in A. platensis. Inositolphosphoceramides (IPC) are minor but important constituents of spirulina; their investigation was accomplished by HILIC-ESI-MS including collision-induced dissociation (MS2 , MS3 ) of deprotonated molecules in the LIT analyzer and a schematic fragmentation pattern is described. All four commercial spirulina samples revealed the occurrence of the same IPC species at m/z 796.6 (d18:0/16:0;1), 810.6 (d18:0/17:0;1), 824.6 (d18:0/18:0;1), and 826.6 (d18:0/17:0;2) but in diverse relative abundance. This study sets the stage for future investigations on IPC in other algae and microalgae.

An easily transferable protocol for in-situ quasi-non-invasive analysis of protein binders in works of art.

Proteomic approaches based on mass spectrometry have become increasingly popular for protein binder's identification in works of art. The identification of the binder employed may offer key information on paintings and other polychrome objects and contribute to assess their historical and technical context, also providing useful hints for a proper restoration and/or conservation treatment. Usually, the protocols employed to this purpose are invasive and at least micro sampling is required. Here, we present a simple transferable method for a quasi-non-invasive analysis of binders in artworks based on the use of a very small poly (2-hydroxyethyl methacrylate)/poly (vinylpyrrolidone) hydrogel (3 mm × 3 mm) previously loaded with trypsin for the in-situ digestion of proteins and applied onto the objects' surface. Upon extraction of digested peptides from the hydrogel, they were examined by MALDI-TOF-MS and/or LC-ESI-MS/MS. The method was validated on fresh and aged model pictorial layers; optical microscope images, and spectrophotocolorimetry confirmed that neither damage nor color alteration of the painting layer occurred, and no hydrogel residue was left. X-ray photoelectron spectroscopy carried out on paint models confirmed that the treatment with trypsin-loaded gels did not modify the pigment composition, even on aged samples. The protocol was successfully applied to a painting on wood mockup aged thirty years, a statue dated XV century exposed in San Lorenzo church (Bisceglie, Bari, Apulia), and a liturgical scroll Benedictio ignis et fontis (Benedizionale) of the Museo Diocesano of Bari dated eleventh century; in all these objects the proteinaceous binder was readily and successfully identified.

MALDI-TOF mass spectrometry analysis of proteins and lipids in Escherichia coli exposed to copper ions and nanoparticles.

Escherichia coli (E. coli) is one of the most important foodborne pathogens to the food industry responsible for diseases as bloody diarrhea, hemorrhagic colitis and life-threatening hemolytic-uremic syndrome. For controlling and eliminating E. coli, metal nano-antimicrobials (NAMs) are frequently used as bioactive systems for applications in food treatments. Most NAMs provide controlled release of metal ions, eventually slowing down or completely inhibiting the growth of undesired microorganisms. Nonetheless, their antimicrobial action is not totally unraveled and is strongly dependent on metal properties and environmental conditions. In this work, we propose the use of matrix-assisted laser desorption ionization time-of-flight (MALDI TOF) mass spectrometry as a powerful tool for direct, time efficient, plausible identification of the cell membrane damage in bacterial strains exposed to copper-based antimicrobial agents, such as soluble salts (chosen as simplified AM material) and copper nanoparticles. E. coli ATCC 25922 strain was selected as 'training bacterium' to set up some critical experimental parameters (i.e. cell concentration, selection of the MALDI matrix, optimal solvent composition, sample preparation method) for the MS analyses. The resulting procedure was then used to attain both protein and lipid fingerprints from E. coli after exposure to different loadings of Cu salts and NPs. Interestingly, bacteria exposed to copper showed over-expression of copper binding proteins and degradation of lipids when treated with soluble salt. These findings were completed with other investigations, such as microbiological experiments. Copyright © 2016 John Wiley & Sons, Ltd.

Superbasic alkyl-substituted bisphosphazene proton sponges: a new class of deprotonating matrices for negative ion matrix-assisted ionization/laser desorption mass spectrometry of low molecular weight hardly ionizable analytes.

RATIONALE: Here hardly ionizable and low molecular weight compounds are detected in negative ion mode by using novel superbasic proton sponges based on 1,8-bisphosphazenylnaphthalene (PN) as MALDI matrices. Among the selected proton sponges, 1,8-bis(trispyrrolidinophosphazenyl)naphthalene (TPPN) has shown the best behaviour as matrix since it allows the direct detection of intact cholesterol without derivatization also in real challenging samples. METHODS: Very weakly acidic compounds such as sterols, steroids, fatty alcohols and saccharides were detected in reflectron negative ion mode by a MALDI TOF/TOF system equipped with a neodymium-doped yttrium lithium fluoride (Nd:YLF) laser (345 nm) with typical mass accuracy of 10 ppm. MS/MS experiments were performed by using ambient air as the collision gas. RESULTS: Contrary to traditional MALDI matrices, superbasic proton sponges allowed the easy deprotonation of an alcohol functional group without a previous chemical derivatization step. Experimental evidence indicates that analyte deprotonation is achieved in the condensed phase, i.e. PN superbasic proton sponges operate according to a recently proposed model named matrix assisted ionization/laser desorption (MAILD). A detection limit of 3 pmol/spot of cholesterol (model compound) with a signal-to-noise ratio ≥ 10 was typically obtained. CONCLUSIONS: For the first time, the usefulness of novel superbasic proton sponges is demonstrated for MALDI detection of hardly ionizable compounds such as sterols, steroids, fatty alcohols and saccharides. The leading candidate TPPN has been successfully applied for negative ion MAILD-MS analysis of cholesterol, fatty acids and phospholipids in egg yolk and brain tissue extracts. Copyright © 2016 John Wiley & Sons, Ltd.

Boronic acid chemistry in MALDI MS: a step forward in designing a reactive matrix with molecular recognition capabilities.

A boronic analogue of the archetype matrix α-cyano-4-hydroxycinnamic acid (CHCA) has been synthesised providing a new "reactive matrix" that possesses molecular recognition properties. This matrix selectively recognizes vic-diols, α-hydroxyacids, aminols and first allowed the detection of anions as fluoride (unaffordable by usual matrices).

1,8-bis(dimethylamino)naphthalene/9-aminoacridine: a new binary matrix for lipid fingerprinting of intact bacteria by matrix assisted laser desorption ionization mass spectrometry.

The effectiveness of a novel binary matrix composed of 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge) and 9-aminoacridine (9AA) for the direct lipid analysis of whole bacterial cells by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) is demonstrated. Deprotonated analyte signals nearly free of matrix-related ions were observed in negative ion mode. The effect of the most important factors (laser energy, pulse voltage, DMAN/9AA ratio, analyte/matrix ratio) was investigated using a Box-Behnken response surface design followed by multi-response optimization in order to simultaneously maximize signal-to-noise (S/N) ratio and resolution. The chemical surface composition of single or mixed matrices was explored by X-ray photoelectron spectroscopy (XPS). Moreover, XPS imaging was used to map the spatial distribution of a model phospholipid in single or binary matrices. The DMAN/9AA binary matrix was then successfully applied to the analysis of intact Gram positive (Lactobacillus sanfranciscensis) or Gram negative (Escherichia coli) microorganisms. About fifty major membrane components (free fatty acids, mono-, di- and tri-glycerides, phospholipids, glycolipids and cardiolipins) were quickly and easily detected over a mass range spanning from ca. 200 to ca. 1600 m/z. Moreover, mass spectra with improved S/N ratio (compared to single matrices), reduced chemical noise and no formation of matrix-clusters were invariably obtained demonstrating the potential of this binary matrix to improve sensitivity.

Fingerprinting of egg and oil binders in painted artworks by matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of lipid oxidation by-products.

A matrix-assisted laser desorption ionization time-of-flight mass spectrometry-based approach was applied for the detection of various lipid classes, such as triacylglycerols (TAGs) and phospholipids (PLs), and their oxidation by-products in extracts of small (50-100 µg) samples obtained from painted artworks. Ageing of test specimens under various conditions, including the presence of different pigments, was preliminarily investigated. During ageing, the TAGs and PLs content decreased, whereas the amount of diglycerides, short-chain oxidative products arising from TAGs and PLs, and oxidized TAGs and PLs components increased. The examination of a series of model paint samples gave a clear indication that specific ions produced by oxidative cleavage of PLs and/or TAGs may be used as markers for egg and drying oil-based binders. Their elemental composition and hypothetical structure are also tentatively proposed. Moreover, the simultaneous presence of egg and oil binders can be easily and unambiguously ascertained through the simultaneous occurrence of the relevant specific markers. The potential of the proposed approach was demonstrated for the first time by the analysis of real samples from a polyptych of Bartolomeo Vivarini (fifteenth century) and a "French school" canvas painting (seventeenth century).

Solid phase microextraction--liquid chromatography (SPME-LC) determination of chloramphenicol in urine and environmental water samples.

A solid phase microextraction--liquid chromatography with ultraviolet detection (SPME-LC-UV) method for the determination of the antimicrobial agent chloramphenicol was developed. The performances of three commercially available fibers were compared; the Carbowax/TPR-100 was found to provide the most efficient extraction. All the aspects influencing the fiber adsorption (extraction time, temperature, pH, salt addition) and desorption (desorption and injection time, desorption solvent mixture composition) of the analyte were investigated. The method was eventually applied to the determination of the drug in both biological (urine) and environmental (tap and sea water) samples. The optimized procedure required a simple sample pretreatment, isocratic elution, and provided enough sensitivity for the analyte determination in the considered samples. The investigated linear ranges were 37-1000 ng/ml (urine), 0.1-10 ng/ml (tap water), 0.3-30 ng/ml (sea water). Within-day and between-days RSD% ranged between 5.5-6.2 and 8.7-9.0 (urine), 5.1-6.0 and 8.4-8.8 (tap water), 5.4-5.7 and 8.6-8.9 (sea water). Estimated LOD and LOQ were 37 and 95 ng/ml (urine), 0.1 and 0.3 ng/ml (tap water), 0.3 and 0.7 ng/ml (sea water).

Optimization of analytical and pre-analytical conditions for MALDI-TOF-MS human urine protein profiles.

Protein analysis in biological fluids, such as urine, by means of mass spectrometry (MS) still suffers for insufficient standardization in protocols for sample collection, storage and preparation. In this work, the influence of these variables on healthy donors human urine protein profiling performed by matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was studied. A screening of various urine sample pre-treatment procedures and different sample deposition approaches on the MALDI target was performed. The influence of urine samples storage time and temperature on spectral profiles was evaluated by means of principal component analysis (PCA). The whole optimized procedure was eventually applied to the MALDI-TOF-MS analysis of human urine samples taken from prostate cancer patients. The best results in terms of detected ions number and abundance in the MS spectra were obtained by using home-made microcolumns packed with hydrophilic-lipophilic balance (HLB) resin as sample pre-treatment method; this procedure was also less expensive and suitable for high throughput analyses. Afterwards, the spin coating approach for sample deposition on the MALDI target plate was optimized, obtaining homogenous and reproducible spots. Then, PCA indicated that low storage temperatures of acidified and centrifuged samples, together with short handling time, allowed to obtain reproducible profiles without artifacts contribution due to experimental conditions. Finally, interesting differences were found by comparing the MALDI-TOF-MS protein profiles of pooled urine samples of healthy donors and prostate cancer patients. The results showed that analytical and pre-analytical variables are crucial for the success of urine analysis, to obtain meaningful and reproducible data, even if the intra-patient variability is very difficult to avoid. It has been proven how pooled urine samples can be an interesting way to make easier the comparison between healthy and pathological samples and to individuate possible differences in the protein expression between the two sets of samples.

Silver nanofractals: electrochemical synthesis, XPS characterization and application in LDI-MS.

Silver nanofractals (Ag-NFs) have been electrosynthesized and characterized by means of morphological and spectroscopic analytical techniques. In particular, X-ray photoelectron spectroscopy has been used to assess the nanomaterial surface chemical state. Ag-NFs show interesting perspectives in bioanalytical applications, particularly as non-conventional desorption and ionization promoters in laser desorption ionization mass spectrometry.

Determination of clenbuterol in human urine and serum by solid-phase microextraction coupled to liquid chromatography.

A solid-phase microextraction (SPME)-LC-UV method for the determination of the beta-adrenergic drug clenbuterol in human urine and serum samples was developed for the first time using a polydimethylsiloxane/divinylbenzene (PDMS/DVB) coated fiber. The procedure required very simple sample pretreatments, isocratic elution, and provided highly selective extractions. All the aspects influencing fiber adsorption (extraction time, temperature, pH, salt addition) and desorption (desorption and injection time, desorption solvent mixture composition) of the analyte have been investigated. The linear ranges investigated in urine and serum were 10-500 and 5-500 ng/ml, respectively (that covers the typical clenbuterol concentration observed in biological fluids). Within-day and between-days R.S.D.% in urine ranged between 5.0-5.3 and 8.5-8.7, respectively, while in serum ranged between 5.5-5.9 and 8.7-9.1, respectively. Estimated LOD and LOQ were 9 and 32 ng/ml (spiked urine), respectively, and 5 and 24 ng/ml (spiked serum), respectively, well below the usual clenbuterol urinary and serum level.

Mouse paracentric inversion In(3)55Rk mutates the urate oxidase gene.

The paracentric inversion In(3)55Rk on mouse Chromosome 3 (Chr 3) was induced by cesium irradiation. Genetic crosses indicate the proximal breakpoint cosegregates with D3Mit324 and D3Mit92; the distal breakpoint cosegregates with D3Mit127, D3Mit160, and D3Mit200. Giemsa-banded chromosomes show the inversion spans approximately 80% of Chr 3. The proximal breakpoint occurs within band 3A2, not 3B as reported previously; the distal breakpoint occurs within band 3H3. Mice homozygous for the inversion exhibit nephropathy indicative of uricase deficiency. Southern blot analyses of urate oxidase, Uox, show two RFLPs of genomic mutant DNA: an EcoRI site between exons 4-8 and a BamHI site 3' to exon 6. Mutant cDNA fails to amplify downstream of base 844 at the 3' end of exon 7. FISH analysis of chromosomes from inversion heterozygotes, using a cosmid clone containing genomic wild-type DNA for Uox exons 2-4, shows that a 5' segment of the mutated Uox allele on the inverted chromosome has been transposed from the distal breakpoint region to the proximal breakpoint region. Clinical, histopathological, and Northern analyses indicate that our radiation-induced mutation, uox(In), is a putative null.

Recurrent endogenous candidal endophthalmitis in a premature infant.

Endogenous Candida endophthalmitis resulting from candidemia in low-birth-weight infants usually occurs as a retinochoroiditis, which is effectively treated with systemic antifungal agents. We report a case of Candida endophthalmitis that recurred 4 months after completion of systemic antifungal therapy. The recurrent Candida infection affected primarily the iris and lens, rather than the retina and choroid. Vitrectomy was required for diagnosis and treatment.

Creation of myelomeningocele in the fetal rabbit.

OBJECTIVE: Myelomeningocele is a neural tube defect resulting in an exposed spinal cord, which leads to irreversible neurologic damage at birth. We proposed development of a fetal rabbit model of myelomeningocele to study in utero spinal cord injury and repair strategies. METHODS: New Zealand white rabbits (n = 10) at 22 days of gestation (term = 31 days) underwent laparotomy to expose the gravid uterus; a hysterotomy exposed the fetal hindlimbs and back. A three to four level lumbar laminectomy was performed, and the dura over the posterior spinal cord was removed. At 30 days of gestation, the does underwent C-section for fetal harvest, and total fetal number, length, weight, and the presence or absence of a spinal defect were recorded for all viable fetuses. RESULTS: All injured fetuses were smaller and weighed less than the nonoperated littermate controls, and histologic examination confirmed a spina bifida-like lesion of their spinal cords. CONCLUSIONS: We successfully created an exposed spinal cord defect in the fetal rabbit model similar to the lesion found in humans. Advantageous because of low animal cost, relatively large fetal size, multiple fetuses per pregnancy, and short total gestation, this model will allow us to study the mechanism of injury to the exposed spinal cord, and perhaps develop strategies to repair human myelomeningoceles.

Tissue removal utilizing Steiner Morcellator within a LapSac: effects of a fluid-filled environment.

BACKGROUND AND PURPOSE: Tissue removal can be a simple process of withdrawal of the entire organ, piecemeal removal with surgical clamps, or mechanical morcellation. Different mechanical morcellators exist that each have advantages and disadvantages. We have investigated a particular morcellator having an internal mechanized blade system that increases the chances of damage to tissue isolation sacks but removes large volumes of intact organ that can more readily be evaluated histologically. The primary premise of this investigation is that a fluid-filled sack would be less likely to be damaged by the activated blades of the morcellator. MATERIALS AND METHODS: Utilizing a Steiner Morcellator (Karl Storz, Culver City, CA), two porcine kidneys were morcellated within the large LapSac (Cook Urological, Spencer, IN). Two environmental variables were evaluated: dry sac morcellation and fluid-filled sac morcellation. Each session was timed, fluid leakage identified, grasping of the sacks quantified, and gross spillage noted. The tissues were submitted for pathologic evaluation to quantify any differences grossly or histologically. All LapSacs were inspected for gross violation and inflated to distention with fluid to check for tiny leaks. RESULTS: The Steiner Morcellator worked much better within the confines of the LapSac filled with fluid. There were no perforations in our experimental setting. It was not possible discern use of fluid-filled sacks histologically. CONCLUSIONS: The Steiner Morcellator can be utilized safely in the LapSac if cautious observation and fluid-filled sack conditions are maintained. The extracted tissue is easily evaluated histologically.

Running suturing: the ideal length facilitates this task.

BACKGROUND AND PURPOSE: Laparoscopic intracorporeal suturing is difficult, a complex task involving several integrated skills such as needle handling, suturing, and knotting. Running suturing is even more complex in the closed environment secondary to the angles of the suture lines, the tension maintained on the suture line, and the need to secure the ends, including tying a knot from the tail of the suture to the loop of the preceding stitch. We have hypothesized that the ideal suture length facilitates this process, and this study was specifically designed to determine the ideal suture length for an intracorporeal running suture. MATERIALS AND METHODS: Latex phantoms were incised for 1 or 2 cm, and each was marked with black points to indicate suture entrance and exit sites. These marks were placed 2 mm from the incision, starting 2 mm proximally and ending 2 mm distally. Suture lengths could then be varied in direct proportion to the length of the incision, keeping all of these points as constants. The ratios of suture length:incision length were 9:1, 10:1, and 11:1. One surgeon performed all of the running suturing utilizing dry 3-0 coated polyglactin with a tapered SH needle. The times needed to create running suture lines were recorded (seconds), the number of ideal entrance and exit points tabulated, and the number of technical errors (missed movements resulting in prolonged suturing) recorded. RESULTS: The lengths of both the incision and the suture affect the ability to perform laparoscopic intracorporeal running suturing. As the incision increased, the suture:incision ratio of 9:1 resulted in the most efficient results (quickest suturing and fewest errors). In smaller suture lines (1 cm), the ratios 10:1 and 11:1 appeared better. CONCLUSIONS: A suture:incision ratio of 9:1 is best for longer suture lines and 10:1 is better for short suture lines.

Laparoscopic augmentation cystoplasty using the novel biomaterial Surgisis: small-intestinal submucosa.

BACKGROUND AND PURPOSE: Urinary bladder augmentation is indicated for diverse conditions, including neurogenic bladder, cancer resection, spinal cord injury, and congenital anomalies. The ideal cystoplasty material is yet to be described. Native gastrointestinal segments commonly used are limited by leakage and small-bowel obstruction, metabolic/nutritional abnormalities, calculi, and malignancy. This study assessed laparoscopic bladder augmentation with porcine small intestinal submucosa (SIS). MATERIALS AND METHODS: Five female pigs (<25 kg) were prepared for surgery under general anesthesia. After Veress needle insufflation, a main 10-mm trocar was placed in the midline for the laparoscope, with two lateral 10-mm ports added for operative instruments. The bladder dome was incised, and a patch of SIS was sewn into the bladder using running 2-0 Vicryl. Three animals served as technical studies. Two additional sows underwent long-term survival surgery: one undiverted and one diverted via a Stamey suprapubic catheter. RESULTS: There were no operative losses. The mean operative time was 140 minutes. The SIS graft held the sutures without tearing. Laparoscopic survey revealed no urine leaks at bladder closure. All five animals voided postoperatively. Urinary extravasation was evident in the three undiverted technique animals. In the other two sows, cystoscopy at 7 days showed intact suture lines without evidence of urinary extravasation and with normal vesicular volumes. Tissue growth was evident, but the graft margins were still discernible. CONCLUSIONS: Laparoscopic bladder augmentation was possible using SIS but at minimal volumes. There were no operative complications; however, the material was difficult to deploy and may benefit from application of an absorbable scaffold. Postoperative urinary drainage is necessary. Further studies will optimize the graft configuration for maximal augmentation.

Experimental study of umbilical cord length as a marker of fetal alcohol syndrome.

Umbilical cord length has long been investigated as a potential marker of intrauterine events that may place the neonate at risk for future adverse developmental sequelae. Experimentally, significantly shortened cords have been reported in association with prenatal exposure to common drugs of abuse. This study in rats reports the time course of effects on umbilical cord length of a daily maternal ethanol gavage (3,200 mg/kg) from gestational day 6 through termination of pregnancy at either day 17, 18, 19, or 20. A total of 786 fetuses derived from 60 litters were examined. Control fetuses demonstrated a linear increase in umbilical cord length and body weight gain during late gestation, findings that support previous studies. The body weights of the ethanol-exposed fetuses were reduced significantly on all gestational days examined, indicating intrauterine growth retardation, a characteristic of fetal alcohol syndrome. Similarly, acute fetal akinesia as well as long-term sequelae stemming from impaired neurological development would result from the elevated blood ethanol levels achieved in this study. The umbilical cords of ethanol-exposed fetuses were significantly shorter on gestational days 19 and 20 in comparison to their controls, while cord lengths on days 17 and 18 were not shortened significantly. A stretch hypothesis has been proposed suggesting that the degree of fetal activity is the main determinant of umbilical cord length. In rats, there is a physiologic diminution of the volume of amniotic fluid (oligohydramnios) in late gestation (day 19 to term), which restricts fetal movements but does not appear to alter the linear relationships between gestational age and cord length in controls, thus arguing against the stretch hypothesis. However, cord lengths in the ethanol-exposed fetuses plateaued in late gestation, suggesting possible adherence to a stretch hypothesis. This dichotomy is discussed emphasizing fetal growth and activity as well as intrauterine space.