Method of magnetic resonance imaging of a sample with ex vivo polarization of an MR imaging agent

The present invention provides a method of magnetic resonance investigation of a sample, preferably of a human or non-human animal body, said method comprising the step of ex vivo polarisation of a high T.sub.1 agent and wherein the polarising agent is optionally seperated from the high T.sub.1 agent before the high T.sub.1 agent is administered to the sample.

FIELD OF THE INVNENTION
 This invention relates to a method of magnetic resonance imaging (MRI).
 BACKGROUND OF THE INVNENTION
 Magnetic resonance imaging (MRI) is a diagnostic technique that has become
 particularly attractive to physicians as it is non-invasive and does not
 involve exposing the patient under study to potentially harmful radiation
 such as X-rays.
 In order to achieve effective contrast between MR images of the different
 tissue types in a subject, it has long been known to administer to the
 subject MR contrast agents (e.g. paramagnetic metal species) which effect
 relaxation times of the MR imaging nuclei in the zones in which they are
 administered or at which they aggregate. Contrast enhancement has also
 been achieved by utilising the "Overhauser effect" in which an esr
 transition in an administered paramagnetic species (hereinafter an OMRI
 contrast agent) is coupled to the nuclear spin system of the imaging
 nuclei. The Overhauser effect (also known as dynamic nuclear polarisation)
 can significantly increase the population difference between excited and
 ground nuclear spin states of selected nuclei and thereby amplify the MR
 signal intensity by a factor of a hundred or more allowing OMRI images to
 be generated rapidly and with relatively low primary magnetic fields. Most
 of the OMRI contrast agents disclosed to date are radicals which are used
 to effect polarisation of imaging nuclei in vivo.
 Techniques are now being developed which involve ex vivo polarisation of
 agents containing MR imaging nuclei, prior to administration and MR signal
 measurement. Such techniques may involve the use of polarising agents, for
 example conventional OMRI contrast agents or hyperpolarised gases to
 achieve ex vivo polarisation of administerable MR imaging nuclei. By
 polarising agent is meant any agent suitable for performing ex vivo
 polarisation of an MR imaging agent.
 The ex vivo method has inter alia the advantage that it is possible to
 avoid administering the whole of, or substantially the whole of, the
 polarising agent to the sample under investigation, whilst still achieving
 the desired polarisation. Thus the method is less constrained by
 physiological factors such as the constraints imposed by the
 administrability, biodegradability and toxicity of OMRI contrast agents in
 in vivo techniques.
 SUMMARY OF THE INVENTION
 It has now been found that ex vivo methods of magnetic resonance imaging
 may be improved by using polarised MR imaging agents comprising nuclei
 capable of emitting magnetic resonance signals in a uniform magnetic field
 (eg MR imaging nuclei such as .sup.13 C or .sup.19 F nuclei) and capable
 of exhibiting a long T.sub.1 relaxation time, preferably additionally a
 long T.sub.2 relaxation time. Such agents will be referred to hereinafter
 as "high T.sub.1 agents". Typically the molecules of a high T.sub.1 agent
 will contain MR imaging nuclei in an amount greater than the natural
 abundance of said nuclei in said molecules (i.e. the agent will be
 enriched with said nuclei).
 Thus viewed from one aspect the present invention provides a method of
 magnetic resonance investigation of a sample, preferably of a human or
 non-human animal body (eg. a mammalian, reptilian or avian body), said
 method comprising:
 (i) subjecting a high T.sub.1 agent to ex vivo polarisation;
 (ii) optionally exposing the high T.sub.1 agent to a uniform magnetic field
 (e.g. the primary field B.sub.o of the imaging apparatus of a weaker field
 e.g. 1 G or more);
 (iii) where step (i) is carried out by means of a polarising agent,
 optionally separating the whole, substantially the whole, or a porion of
 said polarising agent from said high T.sub.1 agent;
 (iv) administering said high T.sub.1 agent to said sample;
 (v) exposing said sample to a second radiation of a frequency selected to
 excite nuclear spin transitions in selected nuclei eg the MR imaging
 nuclei of the high T.sub.1 agent;
 (vi) detecting magnetic resonance signals from said sample; and
 (vii) optionally, generating on image, dynamic flow data, diffusion data,
 perfusion data, physiological data (eg. pH, pO.sub.2, pCO.sub.2,
 temperature or ionic concentrations) or metabolic data from said detected
 signals.
 Thus the invention involves the sequential steps of ex vivo polarisation of
 a high T.sub.1 agent comprising nuclei capable of exhibiting a long
 T.sub.1 relaxation time, administration of the polarised high T.sub.1
 agent (preferably in the absence of a portion of, more preferably
 substantially the whole of, any polarising agent), and conventional in
 vivo MR signal generation and measurement. The MR signals obtained in this
 way may be conveniently converted by conventional manipulations into 2-,
 3- or 4-dimensional data including flow, diffusion, physiological or
 metabolic data.
 Viewed from a further aspect the present invention provides a composition
 comprising a polarised .sup.13 C or .sup.19 F enriched compound together
 with one or more physiologically acceptable cariiers or excipients.
 Viewed from a further aspect the present invention provides a contrast
 medium comprising a polarised high T.sup.1 agent being enriched with
 .sup.13 C nuclei having a T.sub.1 relaxation time of 2 s or more in
 solution at magnetic fields of 0.005-10 T, together with one or more
 physiologically acceptable carriers or excipients.

DETAILED DESCRIPTION OF THE INVENTION
 Suitable high T.sub.1 agents may contain nuclei such as protons. However
 other non-zero nuclear spin nuclei may be useful (eg .sup.19 F, .sup.3 Li,
 .sup.1 H, .sup.13 C, .sup.15 N or .sup.31 P) and .sup.19 F and .sup.13 C
 nuclei are particularly preferred. In this event the MR signals from which
 the image is generated will be substantially only from the high T.sub.1
 agent itself. Nonetheless, where the polarised high T.sub.1 agent is
 present in high concentration in administrable media (eg water), there may
 be significant enough transfer of magnetisation to the water protons to be
 able to perform .sup.1 H-MRI on the water protons of the media. Similarly,
 the polarised high T.sub.1 agent may have a significant enough effect on
 in vivo water protons for conventional .sup.1 H MRI to be carried out on
 those protons.
 Where the MR imaging nuclei is other than a proton (eg .sup.13 C or .sup.19
 F), there will be essentially no interference from background signals (the
 natural abundance of .sup.13 C and .sup.19 F being negible) and image
 contrast will be advantageously high. This is especially true where the
 high T.sub.1 agent itself is enriched above natural abundance. Thus the
 method according to the invention has the benefit of being able to provide
 significant spatial weighting to a generated image. In effect, the
 administration of a polarised high T.sub.1 agent to a selected region of a
 sample (eg by injection) means that the contrast effect may be localised
 to that region. The precise effect of course depends on the extent of
 biodistribution over the period in which the high T.sub.1 agent remains
 significantly polarised. In general, specific body volumes (i.e. regions
 of interest such as the vascular system or specific organs such as the
 brain, kidney, heart or liver) into which the agent is administered may be
 defined with improved signal to noise (particularly improved contrast to
 noise) properties of the resulting images in these volumes.
 In one embodiment, a "native image" of the sample (e.g. body) (ie. one
 obtained prior to administration of the high T.sub.1 agent or one obtained
 for the administered high T.sub.1 agent without prior polarisation as in a
 conventional MR experiment) may be generated to provide structural (eg.
 anatomical) information upon which the image obtained in the method
 according to the invention may be superimposed. A "native image" is
 generally not available where .sup.13 C or .sup.19 F is the imaging
 nucleus because of the low abundance of .sup.13 C and .sup.19 F in the
 body. In this case, a proton MR image may be taken to provide the
 anatomical information upon which the .sup.13 C or .sup.19 F image may be
 superimposed.
 Whilst the high T.sub.1 agent may in general be solid, liquid or gas, it
 should of course by physiologically tolerable or be capable of being
 provided in a physiologically tolerable, administerable form. Preferred
 high T.sub.1 agents are soluble in aqueous media (eg. water) and are of
 course non-toxic where the intended end use is in vivo.
 Conveniently, the high T.sub.1 agent once polarised will remain so for a
 period sufficiently long to allow the imaging procedure to be carried out
 in a comfortable time span. Generally sufficient polarisation will be
 retained by the high T.sub.1 agent in its administerable form (eg. in
 injection solution) if it has a T.sub.1 value (at a field strength of
 0.01-5 T and a temperature in the range 20-40.degree. C.) of at least 2 s,
 preferably at least 5 s, more preferably at least 10 s, especially
 preferably 30 s or longer, more especially perferably 70 s or more, yet
 more especially preferably 100 s or more (for example at 37.degree. C. in
 water at 1 T and a concentraion of at least 1 mM). The high T.sub.1 agent
 may be advantageously an agent with a long T.sub.2 relaxation time.
 The long T.sub.1 relaxation time of certain .sup.13 C nuclei is
 particularly advantageous and certain high T.sub.1 agents containing
 .sup.13 C nuclei are therfore preferred for use in the present method. The
 .gamma.-factor of carbon is about 1/4 of the .gamma.-factor for hydrogen
 resulting in a Larmor frequency of about 10 MHz at 1 T. The rf-absorption
 and reflections in a patient is consequently and advantageously less than
 in water (proton) imaging. Preferably the polarised high T.sub.1 agent has
 an effective .sup.13 C nuclear polarisation corresponding to the one
 obtained at thermal equilibrium at 300 K in a field of 0.1 T or more, more
 preferably 25 T or more, particularly preferably 100 T or more, especially
 preferably 5000 T or more (for example 50 kT). High T.sub.1 agents
 containing .sup.19 F nuclei are also preferred.
 When the electron cloud of a given molecule interacts with atoms in
 surrounding tissue, the shielding of the atom responsible for the the MR
 signal is changed giving rise to a shift in the MR frequency ("the
 chemical shift effect"). When the molecule is metabolised, the chemical
 shift will be changed and high T.sub.1 agents in different chemical
 surroundings may be visualised separately using pulses sensitive to
 chemical shift. When the frequency difference between high T.sub.1
 molecules in different surroundings is 150 Hz or higher (corresponding to
 3.5 ppm or higher at 1 T), the two components may be excited separately
 and visualised in two images. Standard chemical shift selective excitation
 pulses may then be utilised. When the frequency separation is less, the
 two components may not be separated by using frequency selective
 rf-pulses. The phase difference created during the time delay after the
 excitation pulse and before the detection of the MR signal may then be
 used to separate the two components. Phase sensitive imaging pulse
 sequence methods (Dixon, Radiology, 1984, 153: 189-194 and Sepponen, Mag
 Res. Imaging, 3, 163-167, 1985) may be used to generate images visualising
 different chemical surroundings or different metabolites. The long T.sub.2
 relaxation time which may be a characteristic of a high T.sub.1 agent will
 under these circumstances make it possible to use long echo times (TE) and
 still get a high signal to noise ratio. Thus an important advantage of the
 high T.sub.1 agents used in the present method is that they exhibit a
 chemical shift dependent on the local composition of the body in which
 they are localised. Preferred high T.sub.1 agents will exhibit (at 1 T) a
 chemical shift of more than 2 ppm, preferably more than 10 ppm depending
 on whether the high T.sub.1 agent is localised inside or outside the
 vascular system. High T.sub.1 agents containing polarised .sup.13 C nuclei
 (or .sup.19 F nuclei) exhibit large changes in chemical shift in response
 to physiological changes (eg. pH, pO.sub.2, pCO.sub.2, redox potential,
 temperature or ionic concentrations of for example Na.sup.+, K.sup.+,
 Ca.sup.2+) or metabolic activity and therefore may be used to monitor
 these parameters.
 Solid high T.sub.1 agents (e.g. .sup.13 C or .sup.19 F enriched solids) may
 exhibit very long T.sub.1 relaxation times and for this reason are
 especially preferred for use in the present method. The T.sub.1 relaxation
 time may be several hours in the bulk phase, although this may be reduced
 by reduction of grain size and/or addition of paramagnetic impurities eg.
 molecular oxygen. The long relaxation time of solids advantageously allows
 the procedure to be conveniently carried out with less haste and is
 particularly advantageous in allowing the polarised solid high T.sub.1
 agent to be stored or transported prior to pharmaceutical formulation and
 administration. In one embodiment, the polarised high T.sub.1 agent may be
 stored at low temperature eg in frozen form and prior to administration,
 the high T.sub.1 agent may be rapidly warmed to physiological temperatures
 using conventional techniques such as infrared or microwave radiation or
 simply by adding hot, sterile administrable media eg saline.
 For in vivo use, a polarised solid high T.sub.1 agent may be dissolved in
 administrable media (eg water or saline), administered to a subject and
 conventional MR imaging performed. Thus solid high T.sub.1 agents are
 preferably rapidly soluble (eg. water soluble) to assist in formulating
 administrable media. Preferably the high T.sub.1 agent should dissolve in
 a physiologically tolerable carrier (eg water or Ringers solution) to a
 concentration of at least 1 mM at a rate of 1 mM/3 T.sub.1 or more,
 particularly preferably 1 mM/2 T.sub.1 or more, especially preferably 1
 mM/T.sub.1 or more. Where the solid high T.sub.1 agent is frozen, the
 adminstrable medium may be heated, preferably to an extent such that the
 temperature of the medium after mixing is close to 37.degree. C.
 A polarised high T.sub.1 agent may be administered (either alone or with
 additional components such as additional high T.sub.1 agents) in liquid
 form. The retention of polarisation in a liquid medium vis-a-vis a gas
 medium is significantly greater. Thus while T.sub.1 and T.sub.2 are in
 general shorter for the liquid, the T.sub.2 * effect due to diffusion is
 10.sup.5 times less significant for the liquid. Consequently for gaseous
 high T.sub.1 agents the imaging sequence used generally has to be FLASH or
 GRASS while in contrast, more efficient imaging sequences may be used for
 liquids. For example, liquids generally have slower diffusion which makes
 it possible to use sequences such as echo planar imaging (EPI). The
 overall technique will be faster and yield better resolution (voxel size&lt;1
 mm) than conventional techniques (voxel size approx. 1-5 mm) at current
 acquisition times. It will give good images at all fields including in low
 field (eg. 0.01-0.5 T) machines.
 Given that the method of the invention should be carried out within the
 time that the high T.sub.1 agent remains significantly polarised, it is
 desirable for administration of the polarised high T.sub.1 agent to be
 effected rapidly and for the MR measurement to follow shortly thereafter.
 The preferred administration route for the polarised high T.sub.1 agent is
 parental eg by bolus injection, by intravenous, intraarterial or peroral
 injection. The injection time should be equivalent to 5 T.sub.1 or less,
 preferably 3 T.sub.1 or less, particularly preferably T.sub.1 or less,
 especially 0.1 T.sub.1 or less. The lungs may be imaged by spray, eg by
 aerosol spray.
 The high T.sub.1 agent should be preferably enriched with nuclei (eg.
 .sup.19 F and/or .sup.13 C nuclei) having a long T.sub.1 relaxation time.
 Preferred are .sup.13 C enriched high T.sub.1 agents having .sup.13 C at
 one particular position (or more than one particular position) in an
 amount in excess of the natural abundance ie above about 1%. Preferably
 such a single carbon position will have 5% or more .sup.13 C, particularly
 preferably 10% or more, especially preferably 25% or more, more especially
 preferably 50% or more, even more preferably in excess of 99% (e.g.
 99.9%). The .sup.13 C nuclei should preferably amount to &gt;2% of all carbon
 atoms in the compound. The high T.sub.1 agent is preferably .sup.13 C
 enriched at one or more carbonyl or quarternary carbon positions, given
 that a .sup.13 C nuceus in a carbonyl group or in certain quaternary
 carbons may have a T.sub.1 relaxation time typically of more than 2 s,
 preferably more than 5 s, especially preferably more than 30 s. Preferably
 the .sup.13 C enriched compound should be deuterium labelled, especially
 adjacent the .sup.13 C nucleus.
 Preferred .sup.13 C enriched compounds are those in which the .sup.13 C
 nucleus is surronded by one or more non-MR active nuclei such as O, S, C
 or a double bond. Specifically preferred .sup.13 C enriched agents are
 .sup.13 CO.sub.3.sup.2- and H.sup.13 CO.sub.3.sup.- (sodium salt for
 injection and calcium or potassium salt for polarisation).
 Also preferred are the following types of compound (* denotes .sup.13 C
 enriched positions):
 (1) carboxyl compounds comprising 1 to 4 carboxyl groups:
 ##STR1##
 (wherein R represents any straight or branched chain hydrocarbon moiety,
 preferably a highly substituted carbon atom, especially preferably a
 quaternary carbon) and esters, isomers, especially stereoisomers and
 rotamers, thereof:
 (2) substituted mono and biaryl compounds:
 ##STR2##
 (wherein each group R or R' is independently a hydrogen atom, an iodine
 atom, a .sup.19 F atom or a hydrophilic moiety M being any of the
 non-ionizing groups conventionally used to enhance water solubility within
 the field of triiodophenyl X-ray contrast agnets including for example a
 straight chain or branched C.sub.1--10 -alkyl group, preferably a
 C.sub.1--5 group, optionally with one or more CH.sub.2 or CH moieties
 replaced by oxygen or nitrogen atoms and optionally substituted by one or
 more groups selected from oxo, hydroxy, amino, carboxyl derivative, and
 oxo substituted sulphur and phosporus atoms).
 Particular examples of group M include polyhydroxyalkyl, hydroxyalkoxyalkyl
 and hydroxypolyalkoxyalkyl and such groups attached to the phenyl group
 via an amide linkage such as
 hydroxyalkylaminocarbonyl, N-alkyl-hydroxyalkylaminocarbonyl and
 bis-hydroxyalkylaminocarbonyl groups. Preferred among such M groups are
 those containing 1, 2, 3, 4, 5 or 6, especially 1, 2 or 3, hydroxy groups,
 e.g.
 --CONH--CH.sub.2 CH.sub.2 OH
 --CONH--CH.sub.2 CHOHCH.sub.2 OH
 --CONH--CH(CH.sub.2 OH).sub.2
 --CON(CH.sub.2 CH.sub.2 OH).sub.2
 as well as other groups such as
 --CONH.sub.2
 --CONHCH.sub.3
 --OCOCH.sub.3
 --N(COCH.sub.3)H
 --N(COCH.sub.3)C.sub.1--3 -alkyl
 --N(COCH.sub.3)-mono, bis or tris-hydroxy C.sub.1--4 -alkyl
 --N(COCH.sub.2 OH)-mono, bis or tris-hydroxy C.sub.1--4 -alkyl
 --N(COCH.sub.2 OH).sub.2
 --CON(CH.sub.2 CHOHCH.sub.2 OH)(CH.sub.2 CH.sub.2 OH)
 --CONH--C(CH.sub.2 OH).sub.3 and
 --CONH--CH(CH.sub.2 OH)(CHOHCH.sub.2 OH).
 In general, the M groups will preferably each comprise a polyhydroxy
 C.sub.1--4 -alkyl group, such as C.sub.1--4 -alkyl groups substituted by
 1, 2, 3 or 4 hydroxy groups (e.g. hydroxymethyl, 2-hydroxyethyl,
 2,3-bishydroxy-propyl, 1,3-bishydroxyprop-2-yl, 2,3,4-trihydroxybutyl, and
 1,2,4-trihydroxybut-2-yl) optionally connected to the phenyl ring via a
 CO, SO or SO.sub.2 group (e.g. COCH.sub.2 OH or SO.sub.2 CH.sub.2 OH).
 Preferred compounds are those in which two or three non-adjacent R groups
 in the or each C.sub.6 R.sub.5 moiety are iodine and at least one, and
 preferably two or three, R groups in the or each C.sub.6 R.sub.5 moiety
 are M or M.sub.1 moieties; each M independently is a non-ionic hydrophilic
 moiety; and each M.sub.1 independently represents a C.sub.1--4 -alkyl
 group substituted by at least one hydroxyl group and optionally linked to
 the phenyl ring via a cabonyl, suphone or sulphoxide group, at least one R
 group, preferably at least two R groups and especially preferably at least
 one R group in the or each C.sub.6 R.sub.5 moiety, being an M.sub.1
 moiety. Especially preferred are the compounds disclosed in WO-A-96/09282.
 (3) sugars:
 ##STR3##
 (4) ketones:
 ##STR4##
 (wherein R and R' are as defined above)
 (5) ureas:
 ##STR5##
 (6) amides:
 ##STR6##
 (7) amino acids:
 ##STR7##
 peptides and proteins labelled in the carbonyl position, particularly those
 known in the art to be useful for targetting tumour cells. Of the
 proteins, albumin is especially preferred. Polymers are also useful,
 particularly those with low toxicity (eg polylysine) and those with many
 carboxyl groups (eg polyglytamic acid). The following amino acids are
 especially preferred:
 ##STR8##
 (8) carbonates:
 ##STR9##
 (9) nucleotides:
 ##STR10##
 (10) tracers:
 ##STR11##
 and (11) compounds such as:
 ##STR12##
 (wherein R.sup.x denotes any of the conventional side chains suitable for
 use in X-ray contrast agents and A denotes I, D, OR, RC.dbd.O or .sup.19
 F)
 ##STR13##
 In any of the above definitions, useless otherwise specified R, R', R" and
 R"' denote any suitable substituent, preferably a substituent bound by a
 non-magnetic nucleus.
 The partly or wholly deuterated or .sup.19 F analogues of any of these
 compounds are particularly preferred.
 Certain of the above-mentioned .sup.13 C enriched compounds are novel per
 se and form a further aspect of the invention. Compounds which are water
 soluble are particularly preferred.
 In general, .sup.13 C enriched amino acids and any known contrast agents
 from the fields of X-ray contrast agents and MRI contrast agents (the
 chelating agent without the metal counterion eg conventional Gd chelating
 agents without Gd) are preferred as high T.sub.1 agents. Intermediates in
 normal metalbolic cycles such as the citric acid cycle eg. fumaric acid
 and pyruvic acid are preferred for the imaging of metabolic activity.
 T.sub.1 values for .sup.13 C enriched compounds useful in the invention are
 reported in the literature or may be routinely determined. Examples
 include:
 (a) non-water soluble (i.e. soluble in an organic solvent)
 ##STR14##
 (b) water soluble
 ##STR15##
 Ex vivo polarisation may be carried out by any known method and by way of
 example four such methods are described hereinbelow. It is envisaged that,
 in the method according to the invention, the level of polarisation
 achieved should be sufficient to allow the high T.sub.1 agent to achieve a
 diagnostically effective contrast enhancement in the sample to which it is
 subsequently administered in whatever form. In general, it is desirable to
 achieve a level of polarisation which is at least a factor of 2 or more
 above the field in which MRI is performed, preferably a factor of 10 or
 more, particularly preferably 100 or more and especially preferably 1000
 or more, eg. 50 kT.
 In a first embodiment of the method according to the invention, ex vivo
 polarisation of the MR imaging nuclei is effected by an OMRI contrast
 agent. In this embodiment, step (i) of the method comprises:
 (a) bringing an OMRI contrast agent and a high T.sub.1 agent into contact
 in a uniform magnetic field (the primary magnetic field B.sub.o); and
 (b) exposing said OMRI contrast agent to a first radiation of a frequency
 selected to excite electron spin transitions in said OMRI contrast agent.
 It is preferred that the OMRI contrast agent and high T.sub.1 agent are
 present as a composition during polarisation.
 Dynamic nuclear polarisation tray be attained by three possible mechanisms:
 (1) the Overhauser effect, (2) the solid effect and (3) thermal mixing
 effect (see A. Abragam and M. Goldman, Nuclear Magnetism: order and
 disorder, oxford University Press, 1982). The Overhauser effect is a
 relaxation driven process that occurs when the electron-nucleus
 interaction is time-dependent (due to thermal motion or relaxation
 effects) on the time scale of the inverse electron Larmor frequency or
 shorter. Electron-nuclear cross-relaxation results in an exchange of
 energy with the lattice giving rise to an enhanced nuclear polarisation.
 The overall enhancement depends on the relative strength of the scalar and
 dipolar electron-nuclear interaction and the microwave power. For statis
 systems both thermal mixing and the solid effect are operative. In the
 solid effect, the electron spin system is irradiated at a frequency that
 corresponds to the sum or difference of the electronic and nuclear Larmor
 frequencies. The nuclear Zeeman reservoir absorbs or emits the energy
 difference and its spin temperature is modified, resulting in an enhanced
 nuclear polarisation. The efficiency depends on the transition
 probabilities of otherwise forbidden transitions that are allowed due to
 the mixing of nuclear states by non-secular terms of the electron-nuclear
 dipolar interaction. Thermal mixing arises when the electron-electron
 dipolar reservoir establishes thermal contact with the nuclear Zeeman
 reservoirs This takes place when the characteristic electronic resonance
 line width is of the order of the nuclear Larmor frequency.
 Electron-electron cross relaxation between spins with difference in energy
 equal to the nuclear Zeeman energy is absorbed or emitted by the
 electronic dipolar reservoir, changing its spin temperature and the
 nuclear polarisation is enhanced. For thermal mixing both the forbidden
 and the allowed transitions can be involved.
 In the first embodiment where the polarising agent is an OMRI contrast
 agent, the method may be conveniently carried out by using a first magnet
 for providing the polarising magnetic field and a second magnet for
 providing the primary magnetic field for MR imaging. The same magnet could
 be used for both purposes. FIG. 1 of the accompanying drawings is a
 schematic representation of an apparatus suitable for carrying out the
 first embodiment of the invention. A freestanding polarising magnet (1)
 optionally together with a filter surrounds an EPR resonator (2) which
 provides the nuclear polarisation. A container (3) comprising a pump is
 provided for carrying out the contrast composition which is delivered to a
 subject (4) by a delivery line (5). The subject is situated within a
 conventional MR scanner (6).
 In the above apparatus, a dielectric resonator may be used in the dynamic
 nuclear polarisation process. Generally speaking, dynamic nuclear
 polarisation requires a volume with a fairly strong high frequency
 magnetic field and an accompanying electric field which is made as small
 as possible. A dielectric resonator may be used to provide a preferred
 field arrangement in which the magnetic field lines are shaped like a
 straw in a sheaf of corn with an electric field forming circles like the
 thread binding the sheaf. A field arrangement of this type may be formed
 by one of several rings or tubes of a material with a high dielectric
 constant and low loss. The man skilled in the art will appreciate that
 such a tube will exhibit different electromagnetic resonant modes. One of
 the dominant modes has the desired characteristic of electric field
 circulating around the tube axis within the wall and being zero at the
 axis and everywhere perpendicular to it. The magnetic field on the other
 hand is concentrated around the tube axis and mainly directed along it.
 The composition to be polarised is conveniently placed inside the
 resonator which is itself placed inside a metal box with a clearance
 typically of the order of the size of the resonator, and is excited to the
 desired resonance with a coupling loop or the like. The metal box ensures
 that the electromagnetic energy does not leak away by radiation. FIG. 2 of
 the accompanying drawings shows a dielectric resonator (1) (with an axis
 of rotational symmetry (2)) within a metal box (3).
 An alternative to the dielectric resonator is a resonant cavity of Which
 several are known to those skilled in the art. One simple and efficient
 resonant cavity is a metal box, such as a cylindrical metal box. A
 suitable mode is the one known as TM1,1,0 which produces a perpendicular
 magnetic field on the axis of the cavity. It is possible to excite two
 such modes in the same cavity at the same frequency producing fields which
 are mutually perpendicular. By arranging them to have a 90.degree. phase
 difference a rotating field can be produced which is especially efficient
 for implementing dynamic polarisation with a minimum of dissipation in the
 sample. Modes with similar field distributions for different shapes of
 cavities e.g. rectangular cavities are familiar to those skilled in the
 art.
 The composition may also be dispersed into a plurality of compartments
 during the dynamic nuclear polarisation step. Thus the composition might
 be typically divided into parallel channels provided, for example, by
 parallel separating plates, discs or tubes, typically open-ended tubes.
 The electric losses (eddy currents) in the composition caused by the
 magnetic field are decreased by dividing the composition into smaller
 volumes using electrically isolating barriers, preferably situated
 perpendicular to the field. If the composition is in a cylindrical vessel
 surrounded by a dielectric resonator as described hereinbefore, the
 isolating barriers would be planes passing radially from the vessel axis
 to its wall. A simpler and more practical arrangement is to polarise the
 composition in a container which contains a plurality of thin-walled tubes
 of an isolating material such as quartz, glass or plastic. This has the
 advantage of reducing the electric losses in the composition which allows
 a larger volume of composition to be polarised for the same applied
 electromagnetic power. The walls, the inner, outer or both of the tubes
 may similarly serve as the substrate onto which the OMRI contrast agent is
 bound so that pressure applied to one end of the container may force the
 polarized, substantially OMRI contrast agent free, fluid high T.sub.1
 agent from the container, for example with a delivery line leading to the
 subject (patient) undergoing MR examination.
 It is envisaged that in the first embodiment of the method according to the
 invention, use may be made of any known OMRI contrast agent capable of
 polarising a high T.sub.1 agent to an extent such that a diagnostically
 effective contrast enhancement, in the sample to which the high T.sub.1
 agent is administered, is achieved. Where the OMRI contrast agent is a
 paramagnetic free radical, the radical may be conveniently prepared in
 situ from a stable radical precursor by a conventional physical or
 chemical radical generation step shortly before polarisation. This is
 particularly important where the radical has a short half-life. In these
 cases, the radical will normally be non-reusable and may conveniently be
 discarded once the separation step of the method according to the
 invention has been completed.
 In solids, it is preferred to effect dynamic nuclear polarisation by
 irradiating an electron spin at low temperature and high field. Specific
 examples of dynamic nuclear polarisation of solid high T.sub.1 agents are:
 (1) 15N-Ala labelled T4-lysosome and 13C-Glycine in frozen aqueous
 solutions of 60:40 glycerol/water with the free radical 4-amino TEMPO as
 the source of electron polarisation (D. A. Hall, D. Maus, G. Gerfen and R.
 G. Griffin, Science, 19-97), Enhancements of ca.50 and 100 were obtained,
 respectively, at 5T and 40K;
 (2) Carboxy-13C labelled glycine in frozen aqueous solution of 60:40
 glycerol/water with TEMPO as the free radical. An enhancement of 185 at 5T
 and 14K was obtained (G. J. Gerfen, L. R. Becerral D. A. Hall, R. G.
 Griffin, R. J, Temkin, D. J. Singel, J. Chem. Phys. 102(24), 9494-9497
 (1995);
 (3) Dynamic polarisation of protons and deuterons in 1,2-athanedial doped
 with complexes of Cr at 2.5T. The obtained degree of polarisation is 80%
 (W. De Boer and T. O Niinikoski, Nuol. Instrum. Meth. 114, 495 (1974).
 Preferably of course a chosen OMRI contrast agent will exhibit a long
 half-life (preferably at least one hour), long relaxation times (T.sub.1e
 and T.sub.2e), high relaxivity and a small number of ESR transition lines.
 Thus the paramagnetic oxygen-based, sulphur-based or carbon-based organic
 free radicals or magnetic particles, referred to in WO-A-68/10419,
 WO-A-90/00904, WO-A-91/12024, WO-A-93/02711 or WO-A-96/39367 would be
 suitable OMRI contrast agents. A particularly preferred characteristic of
 a chosen OMRI contrast agent is that it exhibits low inherent ESR
 linewidths, preferably less than 500 mG, particularly preferably less than
 400 mG, especially preferably less than 150 mG. Generally speaking,
 organic free radicals such as triarylmethyl and nitroxide radicals provide
 the most likely source of such desirably low linewidths eg. those
 described in WO-A-88/10419, WO-A-90/00904, WO-A-91/12024, WO-A-93/02711 or
 WO-A-96/39367.
 However, OMRI contrast agents useful in the first embodiment of the present
 method are not limited to paramagnetic organic free radicals. Particles
 exhibiting the magnetic properties of paramagnetism, superparamagnetism,
 ferromagnetism or ferrimagnetism may also be useful OMRI contrast agents,
 as may be other particles having associated free electrons.
 Superparamagnatic nanoparticles (eg. iron or iron oxide nanoparticies) may
 be particularly useful. Magnetic particles have the advantages over
 organic free radicals of high stability and a strong electronic/nuclear
 spin coupling (ie. high relaxivity) leading to greater Overhauser
 enhancement factors.
 For the purposes of administration, the high T.sub.1 agent should be
 preferably administered in the absence of the whole of, or substantially
 the whole of, the OMRI contrast agent. Preferably at least 80% of the OMRI
 contrast agent is removed, particularly preferably 90% or more, especially
 preferably 95% or more, most especially 99% or more. In general, it is
 desirable to remove as much OMRI contrast agent as possible prior to
 administration to improve physiological tolerability and to increase
 T.sub.1. Thus preferred OMRI contrast agents for use in the first
 embodiment of the method according to the invention are those which can be
 conveniently and rapidly separated from the polarised high T.sub.1 MR
 imaging agent using known techniques as discussed below. However where the
 OMRI contrast agent is non-toxic, the separation step may be omitted. A
 solid (eg. frozen) composition comprising an OMRI contrast agent and a
 high T.sub.1 agent which has been subjected to polarisation may be rapidly
 dissolved in saline (eg. warm saline) and the mixture injected shortly
 thereafter.
 In the separation step of the first embodiment of the method of the
 invention, it is desirable to remove substantially the whole of the OMRI
 contrast agent from the composition (or at least to reduce it to
 physiologically tolerable levels) as rapidly as possible. Many physical
 and chemical separation or extraction techniques are known in the art and
 may be employed to effect rapid and efficient separation of the OMRI
 contrast agent and high T.sub.1 agent. Clearly the more preferred
 separation techniques are those which can be effected rapidly and
 particularly those which allow separation in less than one second. In this
 respect, magnetic particles (eg. superparamagnetic particles) may be
 advantageously used as the OMRI contrast agent as it will be possible to
 make use of the inherent magnetic properties of the particles to achieve
 rapid separation by known techniques. Similarly, where the OMRI contrast
 agent or the particle is bound to a solid bead, it may be conveniently
 separated from the liquid (i.e. if the solid bead is magnetic by an
 appropriately applied magnetic field).
 for ease of separation of the OMRI contrast agent and the high T.sub.1
 agent, it is particularly preferred that the combination of the two be a
 heterogeneous system, eg. a two phase liquid, a solid in liquid suspension
 or a relatively high surface area solid substrate within a liquid, eg. a
 solid in the form of beads fibres or sheets disposed within a liquid phase
 high T.sub.1 agent. In all cases, the diffusion distance between the high
 T.sub.1 agent and OMRI contrast agent must be small enough to achieve an
 effective Overhauser enhancement. Certain OMRI contrast agents are
 inherently particulate in nature, eg. the paramagnetic particles and
 superparamagnetic agents referred to above. Others may be immobilized on,
 absorbed in or coupled to a solid substrate or support (eg. an organic
 polymer or inorganic matrix such as a zeolite or a silicon material) by
 conventional means. Strong covalent binding between OMRI contrast agent
 and solid substrate or support will, in general, limit the effectiveness
 of the agent in achieving the desired Overhauser effect and so it is
 preferred that the binding, if any, between the OMRI contrast agent and
 the solid support or substrate is weak so that the OMRI contrast agent is
 still capable of free rotation. The OMRI contrast agent may be bound or to
 the to a water insoluble substrate/support prior to the polarisation or
 the OMRI contrast agent may be attached/bound to the substrate/support
 after polarisation. The OMRI contrast agent may then be separated from the
 high T.sub.1 agent e.g. by filtration before administration. The OMRI
 contrast agent may also be bound to a water soluble macromolecule and the
 OMRI contrast agent-macromolecule may be separated from the high T.sub.1
 agent before administration.
 Where the combination of an OMRI contrast agent and high T.sub.1 agent is a
 heterogeneous system, it will be possible to use the different physical
 properties of the phases to carry out separation by conventional
 techniques. For example, where one phase is aqueous and the other
 non-aqueous (solid or liquid) it may be possible to simply decant one
 phase from the other. Alternatively, where the OMRI contrast agent is a
 solid or solid substrate (eg. a bead) suspended in a liquid high T.sub.1
 agent the solid may be separated from the liquid by conventional means eg.
 filtration, gravimetric, chromtographic or centrifugal means. It is also
 envisaged that the OMRI contrast agents may comprise lipophilic moieties
 and so be separated from the high T.sub.1 agent by passage over or through
 a fixed lipophilic medium or the OMRI contrast agent may be chemically
 bound to a lipophilic solid bead. The high T.sub.1 agent may also be in a
 solid (eg. frozen) state during polarisation and in close contact with a
 solid OMRI contrast agent. After polarisation it may be dissolved in
 heated water or saline or melted and removed or separated from the OMRI
 contrast agent where the latter may be toxic and cannot be administered.
 One separation technique makes use of a cation exchange polymer and a
 cationic OMRI contrast agent, eg. a triarylmethyl radical carrying pendant
 carboxylate groups. Alternatively acidifying the solution to around pH 4
 may cause the OMRI contrast agent to precipitate out. Separation may then
 be carried out for example by filtration followed by neutralisation. An
 alternative technique involves adding ions which causes precipitation of
 ionic OMRI agents which may then be filtered off.
 Certain OMRI contrast agents, such as the triarylmethyl radical, may have
 an affinity for proteins. Thus, after polarisation, a composition
 containing an OMRI contrast agent with a protein affinity may be passed
 through or over a protein in a form which exposes a large surface area to
 the agent eg. in particulate or surface bound form. In this way, binding
 of the OMRI contrast agent to the protein enables it to be removed from
 the composition.
 Alternatively when a hydrophilic high T.sub.1 agent is in a solid (eg.
 frozen) form it may be brought into contact with a hydrophobic OMRI
 contrast agent which is dissolved in an organic fluid with a melting
 temperature higher than the high T.sub.1 agent. The mixture is frozen and
 polarisation performed. After polarisation, the mixture is heated and the
 solid OMRI contrast agent and its solvent are removed. The high T.sub.1
 agent will remain hyperpolarised for a significant time in the frozen
 state and may be transported long distances before being dissolved in
 water or saline for injection.
 In a second embodiment of the method according to the invention, ex vivo
 polarisation of the MR imaging nuclei is effected by para-hydrogen
 enriched hydrogen gas. Thus step (i) of the second embodiment of the
 method according to the invention comprises:
 (a) preparing enriched hydrogen;
 (b) reacting said enriched hydrogen with a hydrogenatable high T.sub.1
 agent precursor to produce a hydrogenated high T.sub.1 agent;
 Hydrogen molecules exist in two different forms, namely para hydrogen
 (p-H.sub.2) where the nuclear spins are antiparallel and out of phase (the
 singlet state) and ortho hydrogen (o-H.sub.2) where they are parallel or
 antiparallel and in phase (the triplet state). At room temperature, the
 two forms exist in equilibrium with a 1:3 ratio of para:ortho hydrogen. At
 80K the ratio is 48:52 and at 20K is 99.8:0.2. The rate of equilibration
 is very low in pure hydrogen but in the presence of any of several known
 catalysts (such as Fe.sub.3 O.sub.4 or activated charcoal) an equilibrium
 mixture is rapidly obtained and remains stable at room temperature for
 several hours after liberation from the catalyst. Thus by "enriched
 hydrogen" above is meant hydrogen in which there is a higher than
 equilibrium proportion of para-hydrogen, for example more than 25%,
 preferably 45% or more, more preferably 60% or more, particularly
 preferably 90% or more, especially preferably 99% or more. Typically the
 preparation of enriched hydrogen in step (a) above will be carried out
 catalityically at low temperatures e.g. at 160K or less, preferably at 80K
 or less or more preferably at about 20K.
 Generally speaking, if a para-hydrogen molecule is transferred to a high
 T.sub.1 precursor molecule by means of catalytic hydrogenation (typically
 at elevated pressure (e.g. 50 to 100 bar)), the proton spins remain
 antiparallel and begin to relax to thermal equilibrium with the normal
 time constant T.sub.1 of the hydrogen in the molecule (typically about one
 second). However during relaxation some of the polarisation may be
 transferred to neighbouring nuclei by cross-relaxation or other types of
 coupling. The presence of, for example, a .sup.13 C nucleus with a
 suitable substitution pattern close to the relaxing hydrogen may lead to
 the polarisation being conveniently trapped in the slowly relaxing .sup.13
 C nucleus. An enhancement factor or 2580 has been reported in the
 literature (Barkemeyer et al, 1995 J Am Chem Soc 117, 2927-2928).
 High T.sub.1 agent precursors suitable for use in the second embodiment of
 the present invention are hydrogenatable and will typically possess one or
 more unsaturated bonds, e.g. double or triple carbon-carbon bonds.
 Preferably the high T.sub.1 agent precursor may be .sup.13 C enriched in
 positions close to the hydrogenation site, e.g. a double or triple bond
 where relaxation is slow.
 Generally speaking, to increase the MR signal from the hydrogenated high
 T.sub.1 agent, it is desirable to incorporate more than one unsaturated
 bond in each molecule of the hydragenatable high T.sub.1 agent precursor,
 preferably in a conjugated unsaturated system. However due consideration
 must be given to the need to keep molecular weight relatively low to
 prevent difficulties in administration of the agent. The presence of one
 or more acetylene groups in the hydrogenatable high T.sub.1 agent
 precursor increases the reaction rate and is therefore preferred. More
 preferred are compounds with an unsaturated carbon-carbon bond with one
 ore more carbonyl substituents, e.g. an .alpha..beta. unsaturated carbonly
 compound. Particularly preferred are compounds comprising disubstituted
 unsymmetric alkene or acetylene groups with a
 carbonyl-unsaturation-carbonyl moiety. Such compounds are of high
 reactivity and may allow two or more .sup.13 C atoms to be incorporated to
 utilize the polarisation more efficiently.
 Specifically preferred hydrogenatable high T.sub.1 agent precursors for use
 in the second embodiment of the method of the invention include simple
 acids (e.g. acrylic acid, crotonic acid, propionic acid, furaric acid and
 maleic acid),
 ##STR16##
 quaternary .sup.13 C containing compounds such as
 ##STR17##
 compounds with more than one hydrogenation site such as
 ##STR18##
 and other compounds such as:
 ##STR19##
 and
 ##STR20##
 (where R.sub.1 is
 ##STR21##
 and R is CONHR.sub.2 and R.sub.2 is a conventional hydrophilic group known
 to be useful in X-ray contrast media such as one of the examples given
 hereinbefore)
 Due to their biotolerablity, compounds with quaternary carbons are
 preferred. Cationic compounds may also be used e.g. quaternary ammonium
 salts.
 One especially preferred hydrogenated high T.sub.1 agent is maleic acid
 dimethyl ester which is the hydrogenation product of acetylene
 dicarboxylic acid dimethyl ester.
 Typically the hydragenatable high T.sub.1 agent precursor will undergo
 hydrogenation in the presence of a suitable catalyst, optionally at
 elevated temperature or pressure. The hydrogenation catalyst need not be a
 homogeneous catalyst but during hydrogenation the entire hydrogen molecule
 should be transferred to the host molecule. Some examples of catalysts
 that are able to fulfil this criterion are shown in Table 1.
 TABLE 1
 Hydrogenation catalysts that transfer
 dihydrogen to a double or triple bind
 Water
 Catalyst Synonym Solubility Comment
 (PPh.sub.3)RhCl Wilkinson's - Active when
 catalyst bound to
 zeolite (12 .ANG.)
 [(NBD)Rh + Cationic
 (Amphos).sub.2 ].sup.3+
 (TPPMS).sub.3 RhCl + Anionic
 (HEXNa).sub.2 RhCl + Anionic
 (OCTNa).sub.2 RhCl + Anionic
 IrCl(CO) (PPh.sub.3).sub.2 Vasca's -
 complex
 ##STR22##
 ##STR23##
 ##STR24##
 ##STR25##
 ##STR26##
 ##STR27##
 ##STR28##
 The reaction mechanism of hydrogenation of ethylene with Wilkinson's
 catalyst is shown by way of example in FIG. 3. The oxidative addition of
 enriched hydrogen to the catalyst is generally an equilibrium step which
 means that certain catalysts will also interconvert p-H.sub.2 and
 o-H.sub.2. It is therefore desirable that the chosen hydrogenatable high
 T.sub.1 agent precursor is highly reactive.
 Hydrogenation may be conveniently but not necessarily performed in aqueous
 media and appropriate catalysts for this use should operate efficiently in
 water and conveniently not facilitate the exchange of hydrogen atoms
 between water and the enriched hydrogen, otherwise the polarisation is
 quickly lost. A water soluble rhodium catalyst is one preferred example.
 In order to facilitate rapid separation of catalyst and hydrogenated high
 T.sub.1 agent after hydrogenation, the catalyst is preferably one which is
 immobilized on a solid material e.g. a polymeric material which allows the
 catalyst-bound solid material to be rapidly filtered off after reaction.
 Known examples useful for the second embodiment of the present method
 include catalysts covalently linked to a support or adsorbed on
 derivatized silica.
 An alternative way to remove catalyst from an aqueous solution is to run
 the reaction in the presence of a water-soluble catalyst (eg a rhodium
 catalyst) which may then be removed by filtration through an ion-exchange
 resin or any other sort of filter that can retain the catalyst and allow
 the product to pass. In the preferred case of a cationic catalyst,
 filtration may be carried out through a cation exchanger. One such
 embodiment makes use of an ion-exchange resin bound cationic complex such
 as [(NBD)Rh(Amphos).sub.2 ].sup.3+. The aqueous solution of an anionic or
 neutral product is obtained in the filtrate. The opposite procedure may of
 course be used for anionic catalysts but these are generally less
 preferred. A neutral catalyst may be separated from the high T.sub.2 agent
 by making use of physical characteristics such as lipophilicity. For
 example, a lipophilic catalyst (e.g. Wilkinson's catalyst) may be used in
 a biphasic system such as water/C18-derivatised silica or even two
 immiscible liquids such as water/heptane.
 Hydrogenation may take place advantageously in a non-aqueous media in which
 the hydrogenation product is insoluble (ie. from which it precipitates).
 The increased T.sub.1 of the solid high T.sub.1 agent allows more time for
 isolation and subsequent dissolution in an administrable medium.
 Hydrogenation may also take place with the high T.sub.1 agent precursor
 being insoluble in non-aqueous media but with a particle size as small as
 possible to increase reactive surface area. The use of non-aqueous media,
 preferably media with non-magnetically active nuclei (eg. CS.sub.2 or
 CO.sub.2 under supercritical conditions) advantageously reduces
 polarisation loss from the polarised high T.sub.1 agent and allows the use
 of an extended range of catalysts.
 In a third embodiment of the method according to the invention, ex vivo
 polarisation of the nuclei is effected by a hyperpolarisable gas. In this
 third embodiment, step (i) of the method according to the invention
 comprises:
 (a) hyperpolarising a hyperpolarisable gas before, during or after
 introducing a high T.sub.1 agent thereto whereby to cause nuclear
 polarization of said high T.sub.1 agent.
 By hyperpolarisable gas is meant a gas with a non-zero spin angular
 momentum capable of undergoing an electron transition to an excited
 electron state and thereafter of decaying back to the ground state.
 Depending on the transition that is optically pumped and the helicity of
 the light a positive or negative spin hyperpolarisation may be achieved
 (up to 100%). Examples of gases suitable for use in the third embodiment
 of the method of the invention include the noble gases He (eg. .sup.3 He
 or .sup.4 He) and Xe (eg. .sup.129 Xe), preferably He, particularly
 preferably .sup.3 He. Alkali metal vapours ay also be used eg. Na, K, Rb,
 Cs vapours. Mixtures of the gases may also be used or the hyperolarisable
 gas may be used in liquid or solid form. The term hyperpolarisable gas
 also covers any gas with non-zero nuclear spin which may be polarised by
 optical pumping and is preferably .sup.129 Xe or .sup.3 He.
 It will be appreciated that in the third embodiment of the invention, the
 hyperpolarised gas may transfer polarisation to the nuclear spin system of
 a high T.sub.1 agent directly or indirectly. Where the high T.sub.1 agent
 is to be polarised indirectly by water vapour, it may be advantageously
 water soluble.
 For the purposes of polarisation according to the third embodiment of the
 invention, the high T.sub.1 agent may be generally in gaseous, liquid or
 solid form. One particularly preferred gaseous high T.sub.1 agent is water
 vapour which is conveniently mixed with a hyperpolarisable gas (eg.
 .sup.129 Xe, .sup.3 He or .sup.4 He) at an elevated temperature to
 maintain the vapour. Generally speaking, the more dense the gaseous
 mixture, the more rapid is the polarisation transfer to the water vapour
 so that it is desirable to have the gas mixture under a pressure typically
 above 3 atmospheres, preferably above 30 atmospheres or even more
 preferably above 300 atmospheres. Indirect polarisation transfer may be
 achieved via an intermediate gas medium, for example water vapour.
 Where the high T.sub.1 agent is polarised whilst in a gaseous state, it is
 convenient (for the purposes of separation from the hyperpolarised gas and
 of administration) to be able to rapidly convert it into a liquid or
 solid. This has the added benefit of significantly increasing T.sub.1.
 Thus where water vapour is used as the high T.sub.1 agent, rapid quenching
 is desirable to condense out polarised water, preferably as ice. Thus
 removing the elevated pressure and temperature imposed on the gas mixture
 will lead to rapid cooling and condensation of polarised water. Further
 rapid cooling is possible by adding, for example, cold saturated salt
 solutions (eg. Ringers Solution at -15.degree. C.) or other cooling
 agents. Yet further cooling is possible by, for example, contacting the
 polarised high T.sub.1 agent with a cold surface.
 Water vapour may be created in situ by heating a water/hyperpolarisable gas
 mixture in a suitable chamber. In this case, the inert nature of noble
 gases is a particular advantage. The volume of water vapour used is
 generally 5 liters or more, preferably 10 liters or more, particularly
 preferably 30 liters or more and especially preferably 60 liters or more.
 In practice, the concentration of noble gas required is relatively low. If
 the gas has only nuclear magnetism the pressure should be more than 3
 atmospheres, preferably 30 atmospheres or more and more preferably 300
 atmospheres or more.
 In a preferred embodiment, a hyperpolarised fluid eg. .sup.129 Xe at
 elevated pressure and/or low temperature is passed through a column of
 solid .sup.13 C enriched and/or .sup.19 F enriched high T.sub.1 agent
 until steady state polarisation of the solid is almost achieved. In
 general any of the above-mentioned .sup.13 C enriched agents may be used.
 In another preferred embodiment, a hyperpolarised gas is
 frozen/crystallised on the solid/frozen surface of a solid high T.sub.1
 agent which has been prepared with as large a surface area as possible.
 The mixture may be transported before warm administrable media (eg.
 saline) is added and physiological temperature reached before injection.
 In order to generate a hyperpolarised gas, the gas is first subjected to a
 discharge or other means of excitation (eg. an appropriate radiofrequency)
 which creates a metastable unpaired electron spin state and is then
 optically (eg. laser) pumped at an appropriate frequency to create
 electron hyperpolarisation. The various methods for achieving this are
 well known to those skilled in the art or are described in inter alia U.S.
 Pat. No. 5,545,396.
 Preferred hyperpolarisable gases for use in the third embodiment of the
 method according to the invention are those which can be conveniently and
 rapidly separated from the polarised high T.sub.1 agent. Noble gases are
 particularly useful given their very low boiling points and inertness.
 Preferably the chosen gas will exhibit a long hyperpolarisability
 half-life (preferably at least 1000s, particularly preferably at least
 4000s and especially preferably 8000s or more).
 A hyperpolarised gas may, if desired, be stored for extended periods of
 time in a hyperpolarised state. This is achieved by maintaining the gas at
 very low temperatures, preferably in a frozen state.
 For ease of separation of the hyperpolarisable gas and the high T.sub.1
 agent, the combination of the two may be advantageously a heterogeneous
 system, eg. the high T.sub.1 agent is a solid or liquid at ambient
 temperatures. In all cases, the diffusion distance between the high
 T.sub.1 agent and gas, fluid or solid must be small enough to achieve an
 effective polarisation.
 In the separation step of the third embodiment of the method of the
 invention, it is desirable to remove substantially the whole of the
 hyperpolarisable gas from the composition (or at least to reduce it to
 physiologically tolerable levels) as rapidly as possible. If desired, the
 gas may be reused which may be an important consideration given the
 expense of noble gases. Many physical and chemical separation or
 extraction techniques known in the art may be employed to effect rapid and
 efficient separation of the hyperpolarisable gas and high T.sub.1 agent.
 Clearly the more preferred separation techniques are those which can be
 effected rapidly and particularly those which allow separation in a
 fraction of the relaxation time T.sub.1 of the high T.sub.1 agent.
 In a fourth embodiment of the method of the invention, ex vivo nuclear
 polarisation of the MR imaging nuclei is effected by the use of a high
 field as described in U.S. Pat. No. 5,479,925 (GEC) and U.S. Pat. No.
 5,617,859 (GEC). U.S. Pat. No. 5,479,925 discloses a method for generating
 MR angiograms in which a contrast agent is passed through a small, high
 field polarising magnet ex vivo, in order to generate a high longitudinal
 magnetisation in the agent prior to its administration to the subject.
 There is however no mention or suggestion of the use of high T.sub.1
 agents to achieve an improved effect.
 Generally speaking, polarisation of an MR imaging nuclei may be achieved by
 thermodynamic equilibration at low temperature and high magnetic field.
 Where the contrast medium to be administered is a solid material (e.g.
 crystalline), it may be introduced into a magnetic field at very low
 temperature. Under these conditions, T.sub.1 is very long (typically many
 hours or months) and consequently it takes an unacceptably long time for
 the medium to reach thermodynamic equilibrium. It has however been
 surprisingly found that, if the contrast medium is exposed to a strong
 variable magnetic gradient, T.sub.1 decreases significantly and
 thermodynamic equilibrium may be achieved in a more convenient period of
 time. Thus if the contrast medium undergoes small movements in the
 gradient field for example by exposure to a magnetic field gradient and
 ultrasound or by relative movement within the gradient field, T.sub.1 will
 drop. When thermodynamic equilibrium is attained, all nuclei in the
 contrast medium will be highly polarised relative to room temperature and
 to normal magnetic fields used in MRI. This procedure has the advantage of
 allowing the contrast medium to be removed from the magnet and transported
 in a "ready-to-use" form to the place where it is to be used. Preferably
 but not essentially transport may take place at a relatively low
 temperature (e.g. at liquid nitrogen temperature). The T.sub.1 of the high
 T.sub.1 solid contrast medium will be long enough to allow transport at
 ambient temperature before use.
 The magnetic field strength used in this fourth embodiment of the invention
 should be as high as possible, preferably &gt;1 T, more preferably 5 T or
 more, especially preferably 15 T or more. The temperature should be very
 low e.g. 100 K or less, preferably 1 K or less, especially preferably 1 mK
 or less.
 Thus viewed from a further aspect the present invention provides a process
 for preparing polarised high T.sub.1 agents, said process comprising:
 (i) subjecting a high T.sub.1 agent to a high magnetic field (e.g. 1 T or
 more) at low temperature (e.g. 100 K or less);
 (ii) exposing the agent to a T.sub.1 shortening effect in order to attain
 thermodynamic equilibrium at said low temperature.
 The T.sub.1 shortening effect may be provided by exposure to a variable
 magnetic field gradient but it may also be achieved by adding magnetic
 material (e.g. paramagnetic, superparamagnetic or ferromagnetic materials)
 to the agent during the period when the agent is exposed to low
 temperature. A suitable T.sub.1 shortening agent is Gd but preferred are
 O.sub.2, NO or NO.sup.s which may be conveniently separated from the high
 T.sub.1 agent before transportation and subsequent use.
 In the fourth embodiment of the invention, both the high T.sub.1 agent and
 the aqueous solvent (eg. water) in which it is dissolved may be polarised.
 This may be carried out at low temperature conveniently in the same
 magnetic field and after mixing the administrable composition should be
 warmed very rapidly prior to administration.
 Thus viewed from a further aspect, the present invention provides an
 administrable composition comprising a polarised high T.sub.1 agent and
 polarised water.
 The high T.sub.1 agents used in the method according to the invention may
 be conveniently formulated with conventional pharmaceutical or veterinary
 carriers or excipients. Formulations manufactured or used according to
 this invention may contain, besides the high T.sub.1 agent, formulation
 aids such as are conventional for therapeutic and diagnostic compositions
 in human or veterinary medicine. Thus the formulation may for example
 include stabilizers, antioxidants, osmolality adjusting agents,
 solubilizing agents, emulsifiers, viscosity enhancers, buffers, etc. The
 formulation may be in forms suitable for parenteral (eg. intravenous or
 intraarterial) or enteral (eg. oral or rectal) application, for example
 for application directly into body cavities having external voidance ducts
 (such as the lungs, the gastrointestinal tract, the bladder and the
 uterus), or for injection or infusion into the cardiovascular system.
 However solutions, suspensions and dispersions in physiological tolerable
 carriers eg. water will generally be preferred.
 For use in in vivo imaging, the formulation, which preferably will be
 substantially isotonic, may conveniently be administered at a
 concentration sufficient to yield a 1 micromolar to 10M concentration of
 the high T.sub.1 agent (or even higher where the high T.sub.1 agent is
 water) in the imaging zone; however the precise concentration and dosage
 will of course depend upon a range of factors such as toxicity, the organ
 targeting ability of the high T.sub.1 agent and the administration route.
 The optimum concentration for the MR imaging agent represents a balance
 between various factors. In general, optimum concentrations would in most
 cases lie in the range 0.1 mM to 10M (or even higher where the high
 T.sub.1 agent is water), preferably more than 10 mM, especially more than
 100 mM. Isotonic solution may be especially preferred. In certain
 circumstances concentrations above 1M are preferred. Where water is the MR
 imaging agent the concentration is approximately 56M. Formulations for
 intravenous or intraarterial administration would preferably contain the
 high T.sub.1 agent in concentrations of 10 mM to 10M (or even higher where
 the high T.sub.1 agent is water), especially more than 50 mM. For bolus
 injection the concentration may conveniently be 0.1 mM to 56M, preferably
 more than 200 mM, more preferably more than 500 mM. In certain
 circumstances, the preferred concentration is above 1M, even more
 preferably above 5M. For water as the MR imaging agent the concentration
 is approximately 56M.
 Parenterally administrable forms should of course be sterile and free from
 physiologically unacceptable agents, and should have low osmolality to
 minimize irritation or other adverse effects upon administration and thus
 the formulation should preferably be isotonic or slightly hypertonic.
 Suitable vehicles include aqueous vehicles customarily used for
 administering parenteral solutions such as Sodium Chloride solution,
 Ringer's solution, Dextrose solution, Dextrose and Sodium Chloride
 solution, Lactated Ringer's solution and other solutions such as are
 described in Remington's Pharmaceutical Sciences, 15th ed., Easton: Mack
 Publishing Co., pp. 1405-1412 and 1461-1487 (1975) and The National
 Formulary XIV, 14th ed. Washington: American Pharmaceutical Association
 (1975). The compositions can contain preservatives, antimicrobial agents,
 buffers and antioxidants conventionally used for parenteral solutions,
 excipients and other additives which are compatible with the high T.sub.1
 agents and which will not interfere with the manufacture, storage or use
 of the products.
 Where the high T.sub.1 agent is to be injected, it may be convenient to
 inject simultaneously at a series of administration sites such that a
 greater proportion of the vascular tree may be visualized before the
 polarization is lost through relaxation.
 The dosages of the high T.sub.1 agent used according to the method of the
 present invention will vary according to the precise nature of the high
 T.sub.1 agents used, of the tissue or organ of interest and of the
 measuring apparatus. Preferably the dosage should be kept as low as
 possible while still achieving a detectable contrast effect. In general,
 the maximum dosage will depend on toxicity constraints.
 The invention is illustrated by the following Examples in a non-limiting
 manner:
 EXAMPLE 1
 A high T.sub.1 agent is placed in a chamber at very low temperature (about
 4 K). Fluent O.sub.2 is added and crystallised on the surface of the high
 T.sub.1 agent. In a separate chamber, frozen H.sub.2 O is subjected to the
 same treatment as the high T.sub.1 agent. Both chambers are placed in a
 strong magnetic field (about 15 T) and the temperature kept low.
 When thermodynamic equilibrium is reached, the temperature is increased to
 about 200 K. The oxygen disappears as a gas. The high T.sub.1 agent and
 the frozen H.sub.2 O are mixed and stored until needed. The temperature is
 increased and the solution comprising polarised high T.sub.1 agent and
 hyperpolarised water is injected.
 EXAMPLE 2
 300 mg of sterile Na.sub.2.sup.13 CO.sub.3 or NaH.sup.13 CO.sub.3 is placed
 inside a 10 ml plastic injection syringe. The gas inside the syringe is
 enriched with &gt;20% oxygen. The syringe is placed inside a magnet (1-20 T)
 at a temperature of about 4 K (0.001-5 K) until thermodynamic equilibrium
 is reached.
 The syringe is removed and transported to the subject located in the MRI
 magnet. 10 ml of sterile Ringers Solution (at 37.degree. C., pH 7.4) is
 aspirated and injected at a rate of 10 ml/sec immediately after the high
 T.sub.1 agent has dissolved. .sup.13 C MRI is performed using a fast pulse
 sequence. T.sub.1 in the blood is about 20 s and the distribution of the
 agent is followed on the MR imager.
 EXAMPLE 3
 To a sample of sodium acetate (1-.sup.13 C) is added .alpha.,
 .gamma.-bisphenyl-.beta.-phenylallyl benzene complex (5% w/w). The
 compounds are milled together to give an intimate mixture, which is
 transferred to a borosilicate glass ampule. This is then repeatedly
 evacuated and filled with helium. The last time a pressure of a 200 mbar
 of helium is left in the ampule, which is then flame sealed.
 The sample is polarized by microwaves (70 GHz) for at least one hour at a
 field of 2.5 T at a temperature of 4.2 K. The progress of the polarization
 process is followed by in situ NMR (fast adiabatic passage). When a
 suitable level of polarization has been reached, the ampule is rapidly
 removed from the polarizer and, while handled in a magnetic field of no
 less than 50 mT, cracked open and the contents are quickly discharged and
 dissolved in warm (40.degree. C.) water.
 Experiment 1: This solution is quickly transferred to a spectrometer and
 .sup.13 C spectrum with enhanced intensity is recorded.
 Experiment 2: The sample solution is inserted into an MRI machine with
 .sup.13 C capability and a picture with enhanced intensity and contrast is
 obtained by a single shot technique.
 Experiment 3: The solution is quickly injected into a rat and a .sup.13 C
 MRI picture with enhanced intensity and contrast is obtained, also in this
 case, by utilization of a single shot technique.
 EXAMPLE 4
 To a sample of sodium bicarbonate --.sup.13 C is added MnCl.sub.2 (5% w/w).
 The compounds are milled together to give an intimate mixture, which is
 transferred to a borosilicate glass ampule. This is then repeatedly
 evacuated an filled with helium. The last time a pressure of a 200 mbar of
 helium is left in the ampule, which is then flame sealed.
 The sample is polarized by microwaves (70 GHz) for at least 1 hour at a
 field of 2.5 T at a temperature of 4.2 K. The progress of the polarization
 process is followed by in situ NMR (fast adiabatic passage). When a
 suitable level of polarization has been reached, the ampule is rapidly
 removed from the polarizer and, while handled in a magnetic field of no
 less than 50 mT, cracked open and the contents are quickly discharged and
 dissolved in warm (40.degree. C.) water.
 Experiment 1: This solution is quickly transferred to a spectrometer and
 .sup.13 C spectrum with enhanced intensity is recorded.
 Experiment 2: The sample solution is inserted into an MRI machine with
 .sup.13 C capability and a picture with enhanced intensity and contrast is
 obtained by a single shot technique.
 Experiment 3: The solution is quickly injected into a rat and a .sup.13 C
 MRI picture with enhanced intensity and contrast is obtained, also in this
 case, by utilization of a single shot technique.