C. Leigh Broadhurst

Brain-specific lipids from marine, lacustrine, or terrestrial food resources: potential impact on early African Homo sapiens ☆

The polyunsaturated fatty acid (PUFA) composition of the mammalian central nervous system is almost wholly composed of two long-chain polyunsaturated fatty acids (LC-PUFA), docosahexaenoic acid (DHA) and arachidonic acid (AA). PUFA are dietarily essential, thus normal infant/neonatal brain, intellectual growth and development cannot be accomplished if they are deficient during pregnancy and lactation. Uniquely in the human species, the fetal brain consumes 70% of the energy delivered to it by mother. DHA and AA are needed to construct placental and fetal tissues for cell membrane growth, structure and function. Contemporary evidence shows that the maternal circulation is depleted of AA and DHA during fetal growth. Sustaining normal adult human brain function also requires LC-PUFA.Homo sapiens is unlikely to have evolved a large, complex, metabolically expensive brain in an environment which did not provide abundant dietary LC-PUFA. Conversion of 18-carbon PUFA from vegetation to AA and DHA is considered quantitatively insufficient due to a combination of high rates of PUFA oxidation for energy, inefficient and rate limited enzymatic conversion and substrate recycling. The littoral marine and lacustrine food chains provide consistently greater amounts of pre-formed LC-PUFA than the terrestrial food chain. Dietary levels of DHA are 2.5-100 fold higher for equivalent weights of marine fish or shellfish vs. lean or fat terrestrial meats. Mammalian brain tissue and bird egg yolks, especially from marine birds, are the richest terrestrial sources of LC-PUFA. However, land animal adipose fats have been linked to vascular disease and mental ill-health, whereas marine lipids have been demonstrated to be protective. At South African Capesites, large shell middens and fish remains are associated with evidence for some of the earliest modern humans. Cape sites dating from 100 to 18 kya cluster within 200 km of the present coast. Evidence of early H. sapiens is also found around the Rift Valley lakes and up the Nile Corridor into the Middle East; in some cases there is an association with the use of littoral resources. Exploitation of river, estuarine, stranded and spawning fish, shellfish and sea bird nestlings and eggs by Homo could have provided essential dietary LC-PUFA for men, women, and children without requiring organized hunting/fishing, or sophisticated social behavior. It is however, predictable from the present evidence that exploitation of this food resource would have provided the advantage in multi-generational brain development which would have made possible the advent of H. sapiens. Restriction to land based foods as postulated by the savannah and other hypotheses would have led to degeneration of the brain and vascular system as happened without exception in all other land based apes and mammals as they evolved larger bodies.

Nickel Localization and Response to Increasing Ni Soil Levels in Leaves of the Ni Hyperaccumulator Alyssum murale

We have previously developed phytoremediation and phytomining technologies employing Alyssum Ni hyperaccumulators to quantitatively extract Ni from soils. Implementation of these technologies requires knowledge of Ni localization patterns for the Alyssum species/ecotypes of interest under realistic growth conditions. We investigated Ni uptake and localization in mature Alyssum murale ‘Kotodesh’ and ‘AJ9ç leaves. Seedlings were grown in potting mix with an increasing series of NiSO4 addition (0, 5, 10, 20, 40, 80 mmol Ni kg−1), NiC4H6O4 addition (0, 5, 10, 30, 60, 90 mmol Ni kg−1), in Ni-contaminated soil from metal refining operations, and serpentine soil. Plants at Ni levels 0, 5, 10, 20 mmolkg−1 and in native soils grew normally. Plants at 40 mmolkg−1 exhibited the onset of phytotoxicity, and 60, 80, and 90 mmolkg−1 were demonstrably phytotoxic, but symptoms of phytotoxicity abated within 6 months. Cryogenic complement fractures were made from frozen hydrated samples. High-resolution scanning electron microscope (SEM) images were taken of one half. The other half was freeze-dried and examined with SEM and semi-quantitative energy dispersive x-ray analysis. Ni was highly concentrated in epidermal cell vacuoles and Ni and S counts showed a positive correlation. Trichome pedicles and the epidermal tissue from which the trichome grows were primary Ni compartments, but Ni was not distributed throughout trichomes. Palisade and spongy mesophyll and guard/substomatal cells contained lesser Ni concentrations but palisade mesophyll was an increasingly important compartment as Ni soil levels increased. Ni was virtually excluded from vascular tissue and trichome rays.

Characterization and structure by NMR and FTIR spectroscopy, and molecular modeling of chromium(III) picolinate and nicotinate complexes utilized for nutritional supplementation

Chromium picolinate (CrPic) and chromium nicotinate preparations (CrNicl, CrNic2, CrNic3) were investigated with tH and 13C NMR, FTIR, and molecular modeling. CrPic is crystalline and bidentately coordinated. Cr-PicA bonding broadens the NMR signal or shifts it so far downfieid that it is not detectable. All CrNic preparations are noncrystalline, and results provide no evidence that nicotinic acid (NicA) is O-coordinated to Cr. The complex colors may be due to O-coordination with 1-120 and~or OH, not NicA. I H NMR spectra of CrNic l have two sets of peaks. One set has a significant 8 with respect to NicA, indicating that NicA is more strongly associated with Cr. CrNicl 13C data show small, uniform 8 with respect to NicA, indicating that strong localized Cr-COOH bonding is unlikely. The magnitude of 8, :3C, and 1H exchange data suggests that limited Cr-N bonding may occur in CrNicl. CrNic2 and CrNic3 show little difference from NicA spectra. FTIR spectra of all CrNic complexes, but not CrPic, PicA, or NicA, show bound OH and~ or HzO. CrNic complexes are probably olates, with Cr and NicA OHpolymerized. CrPic exchanges with CrNicl in DMSO. This exchange may provide a mechanism for the absorption and active transport of Cr in biological systems. []ournal of Inorganic Biochemistry 66, 119-130 (1997) @ 1997 Elsevier Science Inc.

The role of docosahexaenoic and the marine food web as determinants of evolution and hominid brain development: The challenge for human sustainability

Life originated on this planet about 3 billion years ago. For the first 2.5 billion years of life there was ample opportunity for DNA modification. Yet there is no evidence of significant change in life forms during that time. It was not until about 600 million years ago, when the oxygen tension rose to a point where air-breathing life forms became thermodynamically possible, that a major change can be abruptly seen in the fossil record. The sudden appearance of the 32 phyla in the Cambrian fossil record was also associated with the appearance of intracellular detail not seen in previous life forms. That detail was provided by cell membranes made with lipids (membrane fats) as structural essentials. Lipids thus played a major, as yet unrecognised, role as determinants in evolution. The compartmentalisation of intracellular, specialist functions as in the nucleus, mitochondria, reticulo-endothelial system and plasma membrane led to cellular specialisation and then speciation. Thus, not only oxygen but also the marine lipids were drivers in the Cambrian explosion. Docosahexaenoic acid (DHA) (all-cis-docosa-4,7,10,13,16,19-hexaenoic acid, C22:6ω3 or C22:6, n-3, DHA) is a major feature of marine lipids. It requires six oxygen atoms to insert its six double bonds, so it would not have been abundant before oxidative metabolism became plentiful. DHA provided the membrane backbone for the emergence of new photoreceptors that converted photons into electricity, laying the foundation for the evolution of other signalling systems, the nervous system and the brain. Hence, the ω3 DHA from the marine food web must have played a critical role in human evolution. There is also clear evidence from molecular biology that DHA is a determinant of neuronal migration, neurogenesis and the expression of several genes involved in brain growth and function. That same process was essential to the ultimate cerebral expansion in human evolution. There is now incontrovertible support of this hypothesis from fossil evidence of human evolution taking advantage of the marine food web. Lipids are still modifying the present evolutionary phase of our species; their signature is evident in the changing panorama of non-communicable diseases. The most worrying change in disease pattern is the sharp rise in brain disorders, which, in the European Union, has overtaken the cost of all other burdens of ill health at €386 billion for the 25 member states at 2004 prices. In 2007, the UK cost was estimated at £77 billion and confirmed in 2010 at £105 billion – greater than heart disease and cancer combined. The rise in mental ill health is now being globalised. The solution to the rising vascular disorders in the last century and now brain disorders in this century lies in a radical reappraisal of the food system, which last century was focussed on protein and calories, with little attention paid to the requirements of the brain – the very organ that was the determinant of human evolution. With the marine fish catch having plateaued 20 years ago and its sustainability now under threat, a critical aspect of this revision is the development of marine agriculture from estuarine, coastal and oceanic resources. Such action is likely to play a key role in future health and intelligence.

Continuous temperature-dependent Raman spectroscopy of melamine and structural analog detection in milk powder.

Hyperspectral Raman imaging has the potential for rapid screening of solid-phase samples for potential adulterants. We can improve mixture analysis algorithms by defining a temperature range in which the contaminant spectrum changes dramatically and uniquely compared with unadulterated material. Raman spectra were acquired for urea, biuret, cyanuric acid, and melamine (pure and at 1% in dried milk powder) from 50 to 310 °C with a gradient of 1 °C min−1. Adulterants were clearly indentified in the milk powder. Specific frequencies that were mainly associated with ring breathing, stretching, and in-plane deformation shifted with respect to temperature up to 12 cm−1 in all four molecules. Specific frequencies significantly increased/decreased in intensity within narrow temperature ranges independent of whether the amine was mixed in milk. Correlation of Raman and differential scanning calorimetry data identified structural components and vibrational modes, which concur with or trigger phase transitions.

Continuous gradient temperature Raman spectroscopy of N-6DPA and DHA from -100 to 20°C.

One of the great unanswered questions with respect to biological science in general is the absolute necessity of docosahexaenoic acid (DHA, 22:6n-3) in fast signal processing tissues. N-6 docosapentaenoic acid (n-6DPA, 22:5n-6), with just one less double bond, group, is fairly abundant in terrestrial food chains yet cannot substitute for DHA. Gradient temperature Raman spectroscopy (GTRS) applies the temperature gradients utilized in differential scanning calorimetry (DSC) to Raman spectroscopy, providing a straightforward technique to identify molecular rearrangements that occur near and at phase transitions. Herein we apply GTRS and DSC to n-6DPA and DHA from -100 to 20°C. 20Mb three-dimensional data arrays with 0.2°C increments and first/second derivatives allowed complete assignment of solid, liquid and transition state vibrational modes, including low intensity/frequency vibrations that cannot be readily analyzed with conventional Raman. N-6DPA and DHA show significant spectral changes with premelting (-33 and -60°C, respectively) and melting (-27 and -44°C, respectively). The CH2(HCCH)CH2 moieties are not identical in the second half of the DHA and DPA structures. DPA has bending (1450cm-1) over almost the entire temperature range. In contrast, DHA contains major CH2 twisting (1265cm-1) with no noticeable CH2 bending, consistent with a flat helical structure with a small pitch. Further modeling of neuronal membrane phospholipids must take into account torsion present in the DHA structure, which essential in determining whether the lipid chain is configured more parallel or perpendicular to the hydrophilic head group.

Continuous gradient temperature Raman spectroscopy and differential scanning calorimetry of N-3DPA and DHA from −100 to 10 °C

Docosahexaenoic acid (DHA, 22:6n-3) is exclusively utilized in fast signal processing tissues such as retinal, neural and cardiac. N-3 docosapentaenoic acid (n-3DPA, 22:5n-3), with just one less double bond, is also found in the marine food chain yet cannot substitute for DHA. Gradient temperature Raman spectroscopy (GTRS) applies the temperature gradients utilized in differential scanning calorimetry (DSC) to Raman spectroscopy, providing a straightforward technique to identify molecular rearrangements that occur near and at phase transitions. Herein we apply GTRS and both conventional and modulated DSC to n-3DPA and DHA from -100 to 20°C. Three-dimensional data arrays with 0.2°C increments and first derivatives allowed complete assignment of solid, liquid and transition state vibrational modes. Melting temperatures n-3DPA (-45°C) and DHA (-46°C) are similar and show evidence for solid-state phase transitions not seen in n-6DPA (-27°C melt). The C6H2 site is an elastic marker for temperature perturbation of all three lipids, each of which has a distinct three dimensional structure. N-3 DPA shows the spectroscopic signature of saturated fatty acids from C1 to C6. DHA does not have three aliphatic carbons in sequence; n-6DPA does but they occur at the methyl end, and do not yield the characteristic signal. DHA appears to have uniform twisting from C6H2 to C12H2 to C18H2 whereas n-6DPA bends from C12 to C18, centered at C15H2. For n-3DPA, twisting is centered at C6H2 adjacent to the C2-C3-C4-C5 aliphatic moiety. These molecular sites are the most elastic in the solid phase and during premelting.

Continuous gradient temperature Raman spectroscopy from −100 to 40 °C yields new molecular models of arachidonic acid and 2-Arachidonoyl-1-stearoyl-sn-glycero-3-phosphocholine

Despite its biochemical importance, a complete Raman analysis of arachidonic acid (AA, 20:4n-6) has never been reported. Gradient temperature Raman spectroscopy (GTRS) applies the temperature gradients utilized in differential scanning calorimetry (DSC) to Raman spectroscopy, providing a straightforward technique to identify molecular rearrangements that occur near and at phase transitions. Herein we utilize the GTRS technique for AA and 1-18:0, 2-20:4n-6 phosphatidyl choline (AAPC) from cryogenic to mammalian body temperatures. 20Mb three-dimensional data arrays with 0.2°C increments and first/second derivatives allowed complete assignment of solid, liquid and transition state vibrational modes. The AA DSC shows a large exothermic peak at -60°C indicating crystallization or a similar major structural change. No exothermic peak of this magnitude was observed in six other unsaturated lipids (DHA, n-3DPA, n-6DPA, LA, ALA, OA). Melting in AA occurs over a large range: (-60 to -35°C): very large frequency offsets and intensity changes correlate with premelting initiating circa -60°C, followed by melting (-37°C). Novel, unique 3D structures for both molecules reveal that AA is not symmetric as a free fatty acid, and it changes significantly when in the sn-2 phospholipid position. Further, different CH and CH2 sites are unequally elastic and nonequivalent.

Continuous gradient temperature Raman spectroscopy of unsaturated fatty acids: applications for fish and meat lipids and rendered meat source identification

Continuous gradient temperature Raman spectroscopy (GTRS) is a simple, rapid technique for determining the unique structures of fatty acids, triacylglycerols and phospholipids. Improved lipid spectra and vibrational assignments have many applications in food safety and quality. Herein we analyze the residual lipid components in porcine and poultry meat and bone meal (MBM) collected directly from rendering operations. We are developing a rapid throughput GTRS method that requires no special extraction methods or toxic/expensive solvents, and can be adapted to field use. Crude ethanol, methanol and water extracts of pork, poultry samples and 20:80 pork:poultry MBM were investigated from -100 to 80°C. GTRS provides 20 Mb three-dimensional data arrays with 0.2°C increments and graphical first and second derivatives. Comparison of second derivative data showed good reproducibility among samples, with some vibrational modes distinct for either pork or chicken. The 20:80 pork:poultry data showed for the first time that lipid prepared from mixed MBM can be positively identified. Oil from pollack waste was also examined. GTRS and other methods can easily identify pure fishmeal or fishmeal mixed into terrestrial MBM; fishmeal is distinguished by its long-chain polyunsaturated fatty acid content. The analytical challenge is to determine economic or accidental adulteration whereby porcine, bovine or ovine MBM are mixed with each other, or mixed into poultry or fish meals.