Neck Muscles' Responses to Cradle, Cross-cradle and Football Breastfeeding Hold Positions in Nursing Mothers: A Preliminary Study.

Background: Breastfeeding-related Neck Pain (BFNP) is prevalent in nursing mothers and cuts across the utilization of different breastfeeding (BF) hold positions. Biomechanical considerations to highlight ergonomically safe BF positions for the prevention of BFNP have not been previously studied. Aim: This study was designed to compare the electrical activities of selected neck muscles across three breastfeeding hold positions [cradle (C1), cross-cradle (C2), and football (FB)]. Materials and Method: Surface electromyographic analyses of four neck muscles (right and left components of each trapezius and sternocleidomastoid) were performed respectively during three BF trials with different BF hold positions (C1, C2, and FB) in 22 nursing mothers. For all the trials, the mothers nursed from the left breast. Results: Across the three BF trials, the electrical activities of each of the neck muscles did not vary significantly (P > 0.05). Furthermore, the left muscular components showed predominantly higher activities, as compared to their right components. Conclusion: The breastfeeding hold position adopted during BF tasks may not be a determinant of BFNP in nursing mothers. Further studies to ascertain the biomechanical implications of the utilized BF holds are recommended.

A novel cationic cardiolipin analogue for gene delivery.

The optically active R and S isomers of cationic cardiolipin analogues (CCA) were synthesized and evaluated as a liposome based transfection reagent. Both isomers form stable liposomes with mean diameters of about 120 nm without any additional lipid ingredients. No significant change in particle size distribution profile was observed over one-month storage at room temperature (20-25 degrees C). The gel to liquid crystalline phase transition temperature (Tm) of cationic liposomes comprised of both R and S isomers was approximately 2 degrees C, as measured by differential scanning calorimetry (DSC). Both isomers also formed stable liposomes when combined with DOPE. In vitro transfection efficiency of the CCA/DOPE liposomes complexed to plasmid DNA was evaluated using a luciferase reporter gene. Both liposomes composed of R and S isomers of the cationic cardiolipin displayed higher transfection efficiency than commercially available Lipofectin. Further in vivo studies are warranted.

A simple method for determination of surface volume mean diameters of oil in water submicron emulsions.

Plots of the droplet size parameter, alpha = (pi(n)D / lambda) versus the total scattering coefficient or specific turbidity, K, are sinusoidal curves. It was noticed that, along one branch of the K-alpha curve (up to 1.1 microm for relative refractive indices (rr(I)) < 1.2), the wavelength dependence on turbidity; i.e. the extent to which light at different wavelengths is scattered showed a progressive decrease with increase in droplet diameter. It was in fact found that when the logarithm of the relative standard deviations (RSD's) in specific turbidities at four different wavelengths (for a particular droplet diameter and relative refractive index) was plotted as a function of that diameter, a straight line resulted. This relationship provides a valuable new method for estimating the surface volume mean droplet size, D(vs), for submicron emulsions by means of simple spectro-turbidimetry.

Development and testing of an improved parenteral formulation of phenytoin using 2-hydroxypropyl-beta-cyclodextrin.

The objective of this study was to increase the solubility of phenytoin by complexing it with varying concentrations of 2-hydroxypropyl-beta-cyclodextrin (HPBCD) and create an entirely aqueous formulation with a pH significantly closer to physiologic pH (7.4). The phenytoin-HPBCD complexation was characterized using phase-solubility analysis at HPBCD concentrations ranging from 10 to 50% w/v over the pH range of 7.4-11.0. The two most promising formulations, i.e., a formulation consisting of 40% HPBCD at pH 10.4, and a second formulation consisting of 20% HPBCD at pH 11.0, were selected for further study. Both formulations were entirely aqueous and had a significantly decreased pH compared to the original commercial formulation (Parke-Davis, pH 12.0). These formulations also exhibited a significantly decreased tendency to precipitate in vitro. The tissue irritation potential of the 20% w/v HPBCD formulation at pH 11.0 was found to be reduced considerably compared to the commercial injection in a BALB/c mouse model.

Systematic screening of antioxidants for maximum protection against oxidation: an oxygen polarograph study.

The development of a simple method for rapid screening of antioxidants in the preformulation phase of drug development is reported. Using an easily oxidizable drug substance containing a tetrahydroisoquinoline nucleus, the relative antioxidant efficacies was determined by simultaneous measurement of dissolved oxygen depletion and drug disappearance rates in presence and absence of antioxidants by oxygen polarographic and high performance liquid chromatographic (HPLC) methods, respectively. Results showed an inverse correlation between oxygen depletion and drug disappearance rates (R2 > 0.85). In contrast, such a high correlation was not obtained when the standard redox potential of these antioxidants was used as a predictor of drug disappearance rates (R2 > 0.50). The rate at which sodium metabisulfite (BIS) and glutathione depleted dissolved oxygen was reduced in the presence of drug, indicating possible reaction between either of the two antioxidants and drug (OHM-11252). Hence, BIS and glutathione may not be suitable antioxidants for stabilizing OHM-11252 in solution. The oxygen depletion rate constants and percent of intact drug remaining were higher in presence of ascorbic acid (ASC) than BIS. Analysis of the reaction mixtures by HPLC diode array detection showed that the number and percent peak area of non-drug peaks formed from ASC-stabilized drug were lower than for BIS. Hence, ASC provides the drug better protection from oxidative degradation.

Characterization of the complexation of diflunisal with hydroxypropyl-beta-cyclodextrin.

The equilibrium dialysis method was applied to the determination of drug cyclodextrin stability constants using diflunisal and 2-hydroxypropyl-beta-cyclodextrin (HPBCD) as a model system. Analysis of the data showed the existence of a linear Scatchard plot, indicative of the formation of a 1:1 diflunisal:HPBCD complex. The mean complexation constant (Kc) +/- S.D. was 3,892 +/- 360 M(-1). The stoichiometry of the complex was verified using the appropriate mass action law equation. The diflunisal:HPBCD complex was also investigated using titration microcalorimetry. A Kc of 3,394 M(-1) was obtained together with an enthalpy change (deltaH) of -20.76 kJ/mol(-1). The Kc values obtained here using the equilibrium dialysis and microcalorimetric methods were comparable to one reported previously using a potentiometric method (5,564 +/- 56 M(-1)).

Skin pigmentation and pharmacokinetics of melanotan-I in humans.

A comparative pharmacokinetic trial was performed with a superpotent synthetic melanotropic peptide, [Nle4-D-Phe7]-alpha-MSHi-13 (melanotan-I or MT-I) given by three routes of administration. Plasma levels were measured by RIA and tanning was quantiated using serial reflectometry. Doses of 0.16 mgkg-1 were administered intravenously (IV) and orally (PO), and doses from 0.08 to 0.21 mg kg-1 subcutaneously (SC), in a randomized crossover fashion to three male volunteers over five consecutive days for 2 weeks (ten doses). The results indicate that the SC dose is completely bioavailable compared to the IV dose. No detectable drug levels were observed following PO dosing. The plasma half-lives following SC dosing ranged from 0.07 to 0.79 h for the absorption phase and from 0.8 to 1.7 h for the beta-phase. Clearance ranged from 0.12 to 0.19 L kg-1 h-1 and 3.9% or less of the dose was recovered in the urine. Side-effects were minimal, consisting of occasional gastrointestinal upset and facial flushing. Significant tanning of the forehead, arms, and neck was noted following IV or SC dosing. This effect peaked at 1 week following drug administration but was still present 3 weeks after completing the ten-dose regimen. It is concluded that SC administration is an efficacious method of delivering melanotan-I.

An improved method for determination of acid dissociation constants of peptides.

PURPOSE: The method described here enables the proton dissociation constants of several amino acid residues of a peptide to be determined simultaneously in aqueous solution without prior knowledge of the exact concentration of the peptide. METHODS: The method used here employs a non-linear fitting program, the BEST program, or a linear least-squares method in combination with the BEST program. These methods are discussed in detail with an emphasis on the quality of the potentiometric titration data that are obtained. Two representative peptides, one with two proton dissociation constants (Ka1, Ka2) and the other with four proton dissociation constants (Ka1-Ka4) were used to illustrate the advantages and the limitations of these two complementary methods. RESULTS: The pKa values of TVL, a schizophrenia-related tripeptide, were found to be 3.62 (+/- 0.02) and 7.17 (+/- 0.05); the pKa values of ELTLQE, a hexapeptide, were found to be 2.32, 3.77, 4.58 and 7.74. CONCLUSIONS: The methods reported here are generally applicable to a variety of peptides. The possibility of integrating these procedures into a preparative chromatographic system for the "on-line" assessment of the pKa values of peptides during the purification stage is an attractive and novel feature of this method.

High-performance liquid chromatographic assay for Melanotan-1 ([Nle4-DPhe7]alpha-melanocyte-stimulating hormone) in biological matrices.

The overall objective of this research was to develop a sensitive, specific, and stability-indicating HPLC assay for the determination of the [Nle4-DPhe7]alpha-melanocyte-stimulating hormone analog known as Melanotan-1 (MT-1) in biological matrices, i.e., cell culture transport media and human plasma. Separation was accomplished isocratically within 8.0 min using a C8 reversed-phase column. The mobile phase consisted of 0.1 M phosphate buffer-acetonitrile (80:20, v/v) with 18 microliters/l triethylamine at pH 2.50. The flow-rate was 1 ml/min with detection at 214 nm. Standard curves (n = 5) were linear over the concentration range 100-1000 ng/ml. The precision, accuracy, intra- and inter-day variations were good with C.V.s typically within 8.7% for concentrations greater than 100 ng/ml. This method was applied to a study of the transport of MT-1 in the Caco-2 cell monolayer model.

Preformulation studies with melanotan-II: a potential skin cancer chemopreventive peptide.

Melanotan-II (1) is a cyclic heptapeptide analogue of alpha-melanocyte-stimulating hormone (alpha-MSH) which tans the skin and is currently being evaluated for the prevention of sunlight-induced skin cancers. The dissociation constants of 1 were determined using potentiometric titration and ultraviolet spectrophotometry. The pKa1 (histidine) and pKa2 (arginine) were estimated to be 6.54 and 11.72, respectively. The apparent partition coefficient (PC) was measured at three pH values using both n-octanol and isooctane as the nonpolar phase. The PC(octanol) and delta log PC at pH 7.35 were 2.82 and 1.05, respectively. These data, together with the observance of a bioavailability of 4.6% in the rat, indicate that 1 may be a suitable candidate for oral delivery. The data presented here are useful in developing an appropriate dosage form for 1.

A comparison of HPLC and bioassay methods for plasma melanotan-II (MT-II) determination: application to a pharmacokinetic study in rats.

The pharmacokinetic profile of the melanotropic peptide, melanotan-II (MT-II), was determined in rats following a 0.3 mg kg-1 intravenous dose. Regression analysis of the plasma MT-II concentrations determined using HPLC and bioassay methods indicated the existence of a significant linear correlation (r = 0.90, p < 0.001). The plasma concentration versus time plots determined using the two assay methods yielded biphasic disposition profiles that were essentially superimposable. The following pharmacokinetic parameters were assessed from plasma concentration versus time data using both methods: Cmax, AUC, CLs, t1/2 beta, MRT, Vd beta, and Vss. Statistical comparison showed that the parameters measured by each method were not significantly different (at the 0.05 level) except for t1/2 beta, MRT and Vss. The presence of even one aberrant data point in the beta-phase can significantly influence t1/2 beta when only a few data points are available in the beta-phase. Since MRT and Vss were calculated from t1/2 beta it is not surprising that these two parameters also differed between methods.

High-performance liquid chromatographic assay for the alpha-melanotropin[4,10] fragment analogue (Melanotan-II) in rat plasma.

A high-performance liquid chromatographic (HPLC) procedure has been developed for the quantification of Melanotan-II (MT-II), a cyclic heptapeptide which promotes rapid tanning of the skin, in rat plasma. The method involves precipitation of plasma proteins followed by direct-injection HPLC with ultraviolet detection. Calibration curves were linear over the range 100-1000 ng/ml for rat plasma. The method is reproducible and reliable with a detection limit of 50 ng/ml in plasma. Within- and between-day precision and accuracy reported as coefficient of variation and relative error, respectively, were < 7%. The application of the assay was successfully demonstrated by quantifying the concentration of MT-II in rat plasma samples following an intravenous dose of 0.3 mg/kg.